Sulfonylalkanoylamino hydroxyethylamino sulfonamide retroviral protease inhibitors

ABSTRACT

Selected sulfonylalkanoylamino hydroxyethylamine sulfonamide compounds are effective as retroviral protease inhibitors, and in particular as inhibitors of HIV protease. The present invention relates to such retroviral protease inhibitors and, more particularly, relates to selected novel compounds, composition and method for inhibiting retroviral proteases, such as human immunodeficiency virus (HIV) protease, prophylactically preventing retroviral infection or the spread of a retrovirus, and treatment of a retroviral infection.

CROSS-REFERENCE TO RELATED APPLICATIONS

This application is a continuation of U.S. Ser. No. 10/082,123 filedFeb. 26, 2002, now U.S. Pat. No. 6,667,307, which is a continuation ofU.S. Ser. No. 09/672,449 filed Sep. 29, 2000 (now U.S. Pat. No.6,380,188); which is a continuation of Ser. No. 09/411,374 filed Oct. 4,1999 (now U.S. Pat. No. 6,169,085); which is a continuation of Ser. No.08/913,069 filed Dec. 19, 1997 (now U.S. Pat. No. 5,985,870); which wasthe national stage entry of PCT/US96/02682, filed Mar. 7, 1996 whichclaims priority to U.S. Ser. No. 08/478,625, filed Jun. 7, 1995, nowU.S. Pat. No. 5,705,500, which was a continuation-in-part of U.S. Ser.No. 08/401,838, filed Mar. 10, 1995, now abandoned.

BACKGROUND OF THE INVENTION

The present invention relates to retroviral protease inhibitors and,more particularly, relates to novel compounds, composition and methodfor inhibiting retroviral proteases, such as human immunodeficiencyvirus (HIV) protease. This invention, in particular, relates tosulfonylalkanoylamino hydroxyethylamine sulfonamide protease inhibitorcompounds, composition and method for inhibiting retroviral proteases,prophylactically preventing retroviral infection or the spread of aretrovirus, and treatment of a retroviral infection, e.g., an HIVinfection. The subject invention also relates to processes for makingsuch compounds as well as to intermediates useful in such processes.

During the replication cycle of retroviruses, gag and gag-pol genetranscription products are translated as proteins. These proteins aresubsequently processed by a virally encoded protease (or proteinase) toyield viral enzymes and structural proteins of the virus core. Mostcommonly, the gag precursor proteins are processed into the coreproteins and the pol precursor proteins are processed into the viralenzymes, e.g., reverse transcriptase and retroviral protease. It hasbeen shown that correct processing of the precursor proteins by theretroviral protease is necessary for assembly of infectious virons. Forexample, it has been shown that frameshift mutations in the proteaseregion of the pol gene of HIV prevents processing of the gag precursorprotein. It has also been shown through site-directed mutagenesis of anaspartic acid residue in the HIV protease active site that processing ofthe gag precursor protein is prevented. Thus, attempts have been made toinhibit viral replication by inhibiting the action of retroviralproteases.

Retroviral protease inhibition typically involves a transition-statemimetic whereby the retroviral protease is exposed to a mimetic compoundwhich binds (typically in a reversible manner) to the enzyme incompetition with the gag and gag-pol proteins to thereby inhibitspecific processing of structural proteins and the release of retroviralprotease itself. In this manner, retroviral replication proteases can beeffectively inhibited.

Several classes of compounds have been proposed, particularly forinhibition of proteases, such as for inhibition of HIV protease. WO92/08701, WO 93/23368, WO 93/23379, WO 94/04493, WO 94/10136 and WO94/14793 (each of which is incorporated herein by reference in itsentirety) for example describe sulfonylalkanoylamino hydroxyethylamine,sulfonylalkanoylamino hydroxyethylurea, sulfonylalkanoylaminohydroxyethyl sulfonamide and sulfonylalkanoylaminohydroxyethylaminosulfonamide isostere containing retroviral proteaseinhibitors. Other such compounds include hydroxyethylamine isosteres andreduced amide isosteres. See, for example, EP O 346 847; EP O 342,541;Roberts et al, “Rational Design of Peptide-Based Proteinase Inhibitors,”Science, 248, 358 (1990); and Erickson et al, “Design Activity, and 2.8Å Crystal Structure of a C₂ Symmetric Inhibitor Complexed to HIV-1Protease,” Science, 249, 527 (1990). U.S. Pat. No. 5,157,041, WO94/04491, WO 94/04492, WO 94/05639 and U.S. patent application Ser. No.08/294,468, filed Aug. 23, 1994, (each of which is incorporated hereinby reference in its entirety) for example describe hydroxyethylamine,hydroxyethylurea or hydroxyethyl sulfonamide isostere containingretroviral protease inhibitors.

Several classes of compounds are known to be useful as inhibitors of theproteolytic enzyme renin. See, for example, U.S. Pat. No. 4,599,198;U.K. 2,184,730; G.B. 2,209,752; EP O 264 795; G.B. 2,200,115 and U.S.SIR H725. Of these, G.B. 2,200,115, GB 2,209,752, EP O 264,795, U.S. SIRH725 and U.S. Pat. No. 4,599,198 disclose urea-containinghydroxyethylamine renin inhibitors. EP 468 641 discloses renininhibitors and intermediates for the preparation of the inhibitors,which include sulfonamide-containing hydroxyethylamine compounds, suchas3-(t-butoxycarbonyl)amino-cyclohexyl-1-(phenylsulfonyl)amino-2(5)-butanol.G.B. 2,200,115 also discloses sulfamoyl-containing hydroxyethylaminerenin inhibitors, and EP 0264 795 discloses certainsulfonamide-containing hydroxyethylamine renin inhibitors. However, itis known that, although renin and HIV proteases are both classified asaspartyl proteases, compounds which are effective renin inhibitorsgenerally are not predictive for effective HIV protease inhibition.

BRIEF DESCRIPTION OF THE INVENTION

The present invention relates to selected retroviral protease inhibitorcompounds, analogs and pharmaceutically acceptable salts, esters andprodrugs thereof. The subject compounds are characterized assulfonylalkanoylamino hydroxyethylamine sulfonamide inhibitor compounds.The invention compounds advantageously inhibit retroviral proteases,such as human immunodeficiency virus (HIV) protease. Therefore, thisinvention also encompasses pharmaceutical compositions, methods forinhibiting retroviral proceases and methods for treatment or prophylaxisof a retroviral infection, such as an HIV infection. The subjectinvention also relates to processes for making such compounds as well asto intermediates useful in such processes.

DETAILED DESCRIPTION OF THE INVENTION

In accordance with the present invention, there is provided a retroviralprotease inhibiting compound of the formula:

or a pharmaceutically acceptable salt, prodrug or ester thereof, whereinn and t each independently represent 0, 1 or 2; preferably n represents1 and t represents 1 or 2;

-   R¹ represents hydrogen, alkyl, alkenyl, alkynyl, hydroxyalkyl,    alkoxyalkyl, cyanoalkyl, —CH₂CONH₂, —CH₂CH₂CONH₂, —CH₂S(O)₂NH₂,    —CH₂SCH₃, —CH₂S(O)CH₃ or —CH₂S(O)₂CH₃ radicals; preferably, R¹    represents hydrogen, alkyl of 1–5 carbon atoms, alkenyl of 2–5    carbon atoms, alkynyl of 2–5 carbon atoms, hydroxyalkyl of 1–3    carbon atoms, alkoxyalkyl of 1–3 alkyl and 1–3 alkoxy carbon atoms,    cyanoalkyl of 1–3 alkyl carbon atoms, —CH₂CONH₂, —CH₂CH₂CONH₂,    —CH₂S(O)₂NH₂, —CH₂SCH₃, —CH₂S(O)CH₃ or —CH₂S(O)₂CH₃ radicals; more    preferably, R¹ represents hydrogen radical, alkyl radical of 1–3    carbon atoms, alkenyl radical of 2–3 carbon atoms, alkynyl radical    of 2–3 carbon atoms or cyanomethyl radicals; even more preferably,    R¹ represents hydrogen, methyl, ethyl or cyanomethyl radicals; yet    more preferably, R¹ represents methyl or ethyl radicals; and most    preferably, R¹ represents a methyl radical;-   R² represents alkyl, aralkyl, alkylthioalkyl, arylthioalkyl or    cycloalkylalkyl radicals; preferably, R² represents radicals of    alkyl of 1–5 carbon atoms, aralkyl of 1–3 alkyl carbon atoms,    alkylthioalkyl of 1–3 alkyl carbon atoms, arylthioalkyl of 1–3 alkyl    carbon atoms or cycloalkylalkyl of 1–3 alkyl carbon atoms and 3–6    ring member carbon atoms; more preferably, R² represents radicals of    alkyl of 3–5 carbon atoms, arylmethyl, alkylthioalkyl of 1–3 alkyl    carbon atoms, arylthiomethyl or cycloalkylmethyl of 5–6 ring member    carbon atoms radicals; even more preferably, R² represents isobutyl,    n-butyl, CH₃SCH₂CH₂—, benzyl, phenylthiomethyl,    (2-naphthylthio)methyl, 4-methoxy phenylmethyl,    4-hydroxyphenylmethyl, 4-fluorophenylmethyl or cyclohexylmethyl    radicals; even more preferably, R² represents benzyl,    4-fluorophenylmethyl or cyclohexylmethyl radicals; most preferably,    R² represents benzyl;-   R³ represents alkyl, cycloalkyl or cycloalkylalkyl radicals;    preferably, R³ represents radicals of alkyl radical of 1–5 carbon    atoms, cycloalkyl of 5–8 ring members or cycloalkylmethyl radical of    3–6 ring members; more preferably, R³ represents propyl, isoamyl,    isobutyl, butyl, cyclopentylmethyl, cyclohexylmethyl, cyclohexyl or    cycloheptyl radicals; more preferably R³ represents isobutyl or    cyclopentylmethyl radicals;-   R⁴ represents heteroaryl or heterocyclo radicals; preferably, R⁴    represents benzo fused 5 to 6 ring member heteroaryl or benzo fused    5 to 6 ring member heterocyclo radicals; or-   R⁴ represents a radical of the formula

wherein A and B each independently represent O, S, SO or SO₂;preferably, A and B each represent O;

-   R⁶ represents deuterium, alkyl or halogen radicals; preferably, R⁶    represents deuterium, alkyl of 1–5 carbon atoms, fluoro or chloro    radicals; more preferably R⁶ represents deuterium, methyl, ethyl,    propyl, isopropyl or fluoro radicals;-   R⁷ represents hydrogen, deuterium, alkyl or halogen radicals;    preferably, R⁷ represents hydrogen, deuterium, alkyl of 1–3 carbon    atoms, fluoro or chloro radicals; more preferably, R⁷ represents    hydrogen, deuterium, methyl or fluoro radicals; or R⁶ and R⁷ each    independently represent fluoro or chloro radicals; and preferably,    R⁶ and R⁷ each represent a fluoro radical; or-   R⁴ represents a radical of the formula

wherein Z represents O, S or NH; and R⁹ represents a radical of formula

wherein Y represents O, S or NH; X represents a bond, O or NR²¹;

-   R²⁰ represents hydrogen, alkyl, alkenyl, alkynyl, aralkyl,    heteroaralkyl, heterocycloalkyl, aminoalkyl, N-mono-substituted or    N,N-disubstituted aminoalkyl wherein said substituents are alkyl or    aralkyl radicals, carboxyalkyl, alkoxycarbonylalkyl, cyanoalkyl or    hydroxyalkyl radicals; preferably, R²⁰ represents hydrogen, alkyl of    1 to 5 carbon atoms, alkenyl of 2 to 5 carbon atoms, alkynyl of 2 to    5 carbon atoms, aralkyl of 1 to 5 alkyl carbon atoms, heteroaralkyl    of 5 to 6 ring members and 1 to 5 alkyl carbon atoms,    heterocycloalkyl of 5 to 6 ring members and 1 to 5 alkyl carbon    atoms, aminoalkyl of 2 to 5 carbon atoms, N-mono-substituted or    N,N-disubstituted aminoalkyl of 2 to 5 alkyl carbon atoms wherein    said substituents are radicals of alkyl of 1 to 3 carbon atoms,    aralkyl of 1 to 3 alkyl carbon atoms radicals, carboxyalkyl of 1 to    5 carbon atoms, alkoxycarbonylalkyl of 1 to 5 alkyl carbon atoms,    cyanoalkyl of 1 to 5 carbon atoms or hydroxyalkyl of 2 to 5 carbon    atoms; more preferably, R²⁰ represents hydrogen, alkyl of 1 to 5    carbon atoms, phenylalkyl of 1 to 3 alkyl carbon atoms,    heterocycloalkyl of 5 to 6 ring members and 1 to 3 alkyl carbon    atoms, or N-mono-substituted or N,N-disubstituted aminoalkyl of 2 to    3 carbon atoms wherein said substituents are alkyl radicals of 1 to    3 carbon atoms; and most preferably, R²⁰ represents hydrogen,    methyl, ethyl, propyl, isopropyl, isobutyl, benzyl,    2-(1-pyrrolidinyl)ethyl, 2-(1-piperidinyl)ethyl,    2-(1-piperazinyl)ethyl, 2-(4-methylpiperazin-1-yl)ethyl,    2-(1-morpholinyl)ethyl, 2-(1-thiamorpholinyl)ethyl or    2-(N,N-dimethylamino)ethyl radicals;-   R²¹ represents hydrogen or alkyl radicals; preferably, R²¹    represents hydrogen radical or alkyl radical of 1 to 3 carbon atoms;    more preferably, R²¹ represents hydrogen or methyl radicals; and    most preferably, R²¹ represents a hydrogen radical; or-   the radical of formula —NR²⁰R²¹ represents a heterocyclo radical;    preferably, the radical of formula —NR²⁰R²¹ represents a 5 to 6 ring    member heterocyclo radical; more preferably, the radical of formula    —NR²⁰R²¹ represents pyrrolidinyl, piperidinyl, piperazinyl,    4-methylpiperazinyl, 4-benzylpiperazinyl, morpholinyl or    thiamorpholinyl radicals; and-   R²² represents alkyl or R²⁰R²¹N-alkyl radicals; preferably, R²²    represents alkyl or R²⁰R²¹N-alkyl radicals wherein alkyl is 1 to 3    carbon atoms; and more preferably, R²² represents alkyl radical of 1    to 3 carbon atoms; and-   preferably R⁴ represents benzothiazol-5-yl, benzothiazol-6-yl,    2-amino-benzothiazol-5-yl, 2-(methoxycarbonylamino)    benzothiazol-5-yl, 2-amino-benzothiazol-6-yl,    2-(methoxycarbonylamino) benzothiazol-6-yl, 5-benzoxazolyl,    6-benzoxazolyl, 6-benzopyranyl, 3,4-dihydrobenzopyran-6-yl,    7-benzopyranyl, 3,4-dihydrobenzopyran-7-yl,    2,3-dihydrobenzofuran-5-yl, benzofuran-5-yl, 1,3-benzodioxol-5-yl,    2-methyl-1,3-benzodioxol-5-yl, 2,2-dimethyl-1,3-benzodioxol-5-yl,    2,2-dideutero-1,3-benzodioxol-5-yl,    2,2-difluoro-1,3-benzodioxol-5-yl, 1,4-benzodioxan-6-yl,    5-benzimidazolyl, 2-(methoxycarbonylamino)benzimidazol-5-yl,    6-quinolinyl, 7-quinolinyl, 6-isoquinolinyl or 7-isoquinolinyl    radicals; more preferably, R⁴ represents benzothiazol-5-yl,    benzothiazol-6-yl, benzoxazol-5-yl, 2,3-dihydrobenzofuran-5-yl,    benzofuran-5-yl, 1,3-benzodioxol-5-yl,    2-methyl-1,3-benzodioxol-5-yl, 2,2-dimethyl-1,3-benzodioxol-5-yl,    2,2-dideutero-1,3-benzodioxol-5-yl,    2,2-difluoro-1,3-benzodioxol-5-yl, 1,4-benzodioxan-6-yl,    2-(methoxycarbonylamino) benzothiazol-5-yl,    2-(methoxycarbonylamino)benzothiazol-6-yl or    2-(methoxycarbonylamino)benzimidazol-5-yl radicals; and most    preferably, R⁴ represents benzothiazol-5-yl, benzothiazol-6-yl,    2,3-dihydrobenzofuran-5-yl, benzofuran-5-yl, 1,3-benzodioxol-5-yl,    2-methyl-1,3-benzodioxol-5-yl, 2,2-dimethyl-1,3-benzodioxol-5-yl,    2,2-dideutero-1,3-benzodioxol-5-yl,    2,2-difluoro-1,3-benzodioxol-5-yl, 1,4-benzodioxan-6-yl,    2-(methoxycarbonylamino)benzothiazol-6-yl or    2-(methoxycarbonylamino)benzimidazol-5-yl radicals; and-   R⁵ represents an alkyl, alkenyl, alkynyl or aralkyl radicals;    preferably, R⁵ represents an alkyl radical of 1–5 carbon atoms,    alkenyl radical of 2–5 carbon atoms, alkynyl radical of 2–5 carbon    atoms or aryl substituted alkyl of 1–5 carbon atoms; more    preferably, R⁵ represents an alkyl radical of 1–5 carbon atoms,    alkenyl radical of 3–4 carbon atoms, alkynyl radical of 3–4 carbon    atoms or aryl substituted alkyl of 1–4 carbon atoms; even more    preferably, R⁵ represents an alkyl radical of 1–5 carbon atoms or    phenyl substituted alkyl of 2–4 carbon atoms; and most preferably,    R⁵ represents an methyl, ethyl, propyl, isopropyl or 2-phenylethyl    radicals.

Preferably, the absolute stereochemistry of the carbon atom of —CH(OH)—group is (R) and the absolute stereochemistry of the carbon atoms of—CH(R¹)— and —CH(R²)— groups is (S).

A family of compounds of particular interest within Formula I arecompounds embraced by the formula

or a pharmaceutically acceptable salt, prodrug or ester thereof, whereint, R¹, R², R³, R⁴ and R⁵ are as defined above.

A family of compounds of further interest within formula II arecompounds embraced by the formula

or a pharmaceutically acceptable salt, prodrug or ester thereof, whereint, R¹, R², R³, R⁴ and R⁵ are as defined above.

A more preferred family of compounds within Formula III consists ofcompounds or a pharmaceutically acceptable salt, prodrug or esterthereof, wherein t represents 2;

-   R¹ represents methyl or ethyl radicals;-   R² represents a benzyl, 4-fluorophenylmethyl or cyclohexylmethyl    radical;-   R³ represents propyl, isoamyl, isobutyl, butyl, cyclohexyl,    cycloheptyl, cyclopentylmethyl or cyclohexylmethyl radicals;-   R⁴ represents 2,3-dihydrobenzofuran-5-yl, 1,3-benzodioxol-5-yl,    2-methyl-1,3-benzodioxol-5-yl, 2,2-dimethyl-1,3-benzodioxol-5-yl,    benzothiazol-6-yl, 2,2-dideutero-1,3-benzodioxol-5-yl,    2,2-difluoro-1,3-benzodioxol-5-yl or 1,4-benzodioxan-6-yl radicals;    and-   R⁵ represents methyl, ethyl, propyl, isopropyl or 2-phenylethyl    radicals.

Compounds of interest include the following:

-   N-[2R-hydroxy-3-[(2-methylpropyl)[(1,3-benzodioxol-5-yl)    sulfonyl]amino]-1S-(phenylmethyl)propyl]-2S-methyl-3-(methylsulfonyl)propanamide;-   N-[2R-hydroxy-3-[(2-methylpropyl)[(1,4-benzodioxan-6-yl)    sulfonyl]amino]-1S-(phenylmethyl)propyl]-2S-methyl-3-(methylsulfonyl)propanamide;-   N-[2R-hydroxy-3-[(2-methylpropyl)[(benzothiazol-6-yl)    sulfonyl]amino]-1S-(phenylmethyl)propyl]-2S-methyl-3-(methylsulfonyl)propanamide;-   N-[2R-hydroxy-3-[(2-methylpropyl)[(benzothiazol-5-yl)    sulfonyl]amino]-1S-(phenylmethyl)propyl]-2S-methyl-3-(methylsulfonyl)propanamide;    and-   N-[2R-hydroxy-3-[(2-methylpropyl)[(2,3-dihydrobenzofuran-5-yl)sulfonyl]amino]-1S-(phenylmethyl)propyl]-2S-methyl-3-(methylsulfonyl)propanamide.

As utilized herein, the term “alkyl”, alone or in combination, means astraight-chain or branched-chain alkyl radical containing preferablyfrom 1 to 8 carbon atoms, more preferably from 1 to 5 carbon atoms, mostpreferably 1–3 carbon atoms. Examples of such radicals include methyl,ethyl, n-propyl, isopropyl, n-butyl, isobutyl, sec-butyl, tert-butyl,pentyl, iso-amyl, hexyl, octyl and the like. The term “hydroxyalkyl”,alone or in combination, means an alkyl radical as defined above whereinat least one hydrogen radical is replaced with a hydroxyl radical,preferably 1–3 hydrogen radicals are replaced by hydroxyl radicals, morepreferably, 1–2 hydrogen radicals are replaced by hydroxyl radicals, andmost preferably, one hydrogen radical is replaced by a hydroxyl radical.The term “alkenyl”, alone or in combination, means a straight-chain orbranched-chain hydrocarbon radical having one or more double bonds andcontaining preferably from 2 to 8 carbon atoms, more preferably from 2to 5 carbon atoms, most preferably from 2 to 3 carbon atoms. Examples ofsuitable alkenyl radicals include ethenyl, propenyl, 2-methylpropenyl,1,4-butadienyl and the like. The term “alkynyl”, alone or incombination, means a straight-chain or branched chain hydrocarbonradical having one or more triple bonds and containing preferably from 2to 8 carbon atoms, more preferably from 2 to 5 carbon atoms, mostpreferably from 2 to 3 carbon atoms. Examples of alkynyl radicalsinclude ethynyl, propynyl (propargyl), butynyl and the like. The term“alkoxy”, alone or in combination, means an alkyl ether radical whereinthe term alkyl is as defined above. Examples of suitable alkyl etherradicals include methoxy, ethoxy, n-propoxy, isopropoxy, n-butoxy,iso-butoxy, sec-butoxy, tert-butoxy and the like. The term “cycloalkyl”,alone or in combination, means a saturated or partially saturatedmonocyclic, bicyclic or tricyclic alkyl radical wherein each cyclicmoiety contains preferably from 3 to 8 carbon atom ring members, morepreferably from 3 to 7 carbon atom ring members, most preferably from 5to 6 carbon atom ring members, and which may optionally be a benzo fusedring system which is optionally substituted as defined herein withrespect to the definition of aryl. Examples of such cycloalkyl radicalsinclude cyclopropyl, cyclobutyl, cyclopentyl, cyclohexyl, cycloheptyl,octahydronaphthyl, 2,3-dihydro-1H-indenyl, adamantyl and the like.“Bicyclic” and “tricyclic” as used herein are intended to include bothfused ring systems, such as naphthyl and β-carbolinyl, and substitutedring systems, such as biphenyl, phenylpyridyl, naphthyl anddiphenylpiperazinyl. The term “cycloalkylalkyl” means an alkyl radicalas defined above which is substituted by a cycloalkyl radical as definedabove. Examples of such cycloalkylalkyl radicals includecyclopropylmethyl, cyclobutylmethyl, cyclopentylmethyl,cyclohexylmethyl, 1-cyclopentylethyl, 1-cyclohexylethyl,2-cyclopentylethyl, 2-cyclohexylethyl, cyclobutylpropyl,cyclopentylpropyl, cyclohexylbutyl and the like. The term “benzo”, aloneor in combination, means the divalent radical C₆H₄═ derived frombenzene. The term “aryl”, alone or in combination, means a phenyl ornaphthyl radical which is optionally substituted with one or moresubstituents selected from alkyl, alkoxy, halogen, hydroxy, amino,nitro, cyano, haloalkyl, carboxy, alkoxycarbonyl, cycloalkyl,heterocyclo, alkanoylamino, amido, amidino, alkoxycarbonylamino,N-alkylamidino, alkylamino, dialkylamino, N-alkylamido,N,N-dialkylamido, aralkoxycarbonylamino, alkylthio, alkylsulfinyl,alkylsulfonyl and the like. Examples of aryl radicals are phenyl,p-tolyl, 4-methoxyphenyl, 4-(tert-butoxy)phenyl,3-methyl-4-methoxyphenyl, 4-CF₃-phenyl, 4-fluorophenyl, 4-chlorophenyl,3-nitrophenyl, 3-aminophenyl, 3-acetamidophenyl, 4-acetamidophenyl,2-methyl-3-acetamidophenyl, 2-methyl-3-aminophenyl,3-methyl-4-aminophenyl, 2-amino-3-methylphenyl,2,4-dimethyl-3-aminophenyl, 4-hydroxyphenyl, 3-methyl-4-hydroxyphenyl,1-naphthyl, 2-naphthyl, 3-amino-1-naphthyl, 2-methyl-3-amino-1-naphthyl,6-amino-2-naphthyl, 4,6-dimethoxy-2-naphthyl, piperazinylphenyl and thelike. The terms “aralkyl” and “aralkoxy”, alone or in combination, meansan alkyl or alkoxy radical as defined above in which at least onehydrogen atom is replaced by an aryl radical as defined above, such asbenzyl, benzyloxy, 2-phenylethyl, dibenzylmethyl, hydroxyphenylmethyl,methylphenylmethyl, diphenylmethyl, diphenylmethoxy,4-methoxyphenylmethoxy and the like. The term “aralkoxycarbonyl”, aloneor in combination, means a radical of the formula aralkyl-O—C(O)— inwhich the term “aralkyl” has the significance given above. Examples ofan aralkoxycarbonyl radical are benzyloxycarbonyl and4-methoxyphenylmethoxycarbonyl. The term “aryloxy” means a radical ofthe formula aryl-O— in which the term aryl has the significance givenabove. The term “alkanoyl”, alone or in combination, means an acylradical derived from an alkanecarboxylic acid, examples of which includeacetyl, propionyl, butyryl, valeryl, 4-methylvaleryl, and the like. Theterm “cycloalkylcarbonyl” means an acyl radical of the formulacycloalkyl-C(O)— in which the term “cycloalkyl” has the significancegive above, such as cyclopropylcarbonyl, cyclohexylcarbonyl,adamantylcarbonyl, 1,2,3,4-tetrahydro-2-naphthoyl,2-acetamido-1,2,3,4-tetrahydro-2-naphthoyl,1-hydroxy-1,2,3,4-tetrahydro-6-naphthoyl and the like. The term“aralkanoyl” means an acyl radical derived from an aryl-substitutedalkanecarboxylic acid such as phenylacetyl, 3-phenylpropionyl(hydrocinnamoyl), 4-phenylbutyryl, (2-naphthyl)acetyl,4-chlorohydrocinnamoyl, 4-aminohydrocinnamoyl, 4-methoxyhydrocinnamoyl,and the like. The term “aroyl” means an acyl radical derived from anarylcarboxylic acid, “aryl” having the meaning given above. Examples ofsuch aroyl radicals include substituted and unsubstituted benzoyl ornapthoyl such as benzoyl, 4-chlorobenzoyl, 4-carboxybenzoyl,4-(benzyloxycarbonyl)benzoyl, 1-naphthoyl, 2-naphthoyl, 6-carboxy-2naphthoyl, 6-(benzyloxycarbonyl)-2-naphthoyl, 3-benzyloxy-2-naphthoyl,3-hydroxy-2-naphthoyl, 3-(benzyloxyformamido)-2-naphthoyl, and the like.The terms “heterocyclo,” alone or in combination, means a saturated orpartially unsaturated monocyclic, bicyclic or tricyclic heterocycleradical containing at least one, preferably 1 to 4, more preferably 1 to2, nitrogen, oxygen or sulfur atom ring member and having preferably 3to 8 ring members in each ring, more preferably 3 to 7 ring members ineach ring and most preferably 5 to 6 ring members in each ring.“Heterocyclo” is intended to include sulfones, sulfoxides, N-oxides oftertiary nitrogen ring members, and carbocyclic fused and benzo fusedring systems. Such heterocyclo radicals may be optionally substituted onat least one, preferably 1 to 4, more preferably 1 to 2, carbon atoms byhalogen, alkyl, alkoxy, hydroxy, oxo, aryl, aralkyl, heteroaryl,heteroaralkyl, amidino, N-alkylamidino, alkoxycarbonylamino,alkylsulfonylamino and the like, and/or on a secondary nitrogen atom(i.e., —NH—) by hydroxy, alkyl, aralkoxycarbonyl, alkanoyl,heteroaralkyl, phenyl or phenylalkyl and/or on a tertiary nitrogen atom(i.e., ═N—) by oxido. “Heterocycloalkyl” means an alkyl radical asdefined above in which at least one hydrogen atom is replaced by aheterocyclo radical as defined above, such as pyrrolidinylmethyl,tetrahydrothienylmethyl, pyridylmethyl and the like. The term“heteroaryl”, alone or in combination, means an aromatic heterocycloradical as defined above, which is optionally substituted as definedabove with respect to the definitions of aryl and heterocyclo. Examplesof such heterocyclo and heteroaryl groups are pyrrolidinyl, piperidinyl,piperazinyl, morpholinyl, thiamorpholinyl, pyrrolyl, imidazolyl (e.g.,imidazol 4-yl, 1-benzyloxycarbonylimidazol-4-yl, etc.), pyrazolyl,pyridyl, (e.g., 2-(1-piperidinyl)pyridyl and 2-(4-benzylpiperazin-1-yl-1-pyridinyl, etc.), pyrazinyl, pyrimidinyl, furyl,tetrahydrofuryl, thienyl, tetrahydrothienyl and its sulfoxide andsulfone derivatives, triazolyl, oxazolyl, thiazolyl, indolyl (e.g.,2-indolyl, etc.), quinolinyl, (e.g., 2-quinolinyl, 3-quinolinyl,1-oxido-2-quinolinyl, etc.), isoquinolinyl (e.g., 1-isoquinolinyl,3-isoquinolinyl, etc.), tetrahydroquinolinyl (e.g.,1,2,3,4-tetrahydro-2-quinolyl, etc.), 1,2,3,4-tetrahydroisoquinolinyl(e.g., 1,2,3,4-tetrahydro-1-oxo-isoquinolinyl, etc.), quinoxalinyl,β-carbolinyl, 2-benzofurancarbonyl, 1-,2-,4- or 5-benzimidazolyl,methylenedioxyphen-4-yl, methylenedioxyphen-5-yl, ethylenedioxyphenyl,benzothiazolyl, benzopyranyl, benzofuryl, 2,3-dihydrobenzofuryl,benzoxazolyl, thiophenyl and the like. The term “heteroaralkyl”, aloneor in combination, means an alkyl radical as defined above in which atleast one hydrogen atom is replaced by an heteroaryl radical as definedabove, such as benzofurylmethyl, 3-furylpropyl, quinolinylmethyl,2-thienylethyl, pyridylmethyl, 2-pyrrolylpropyl, 1-imidazolylethyl andthe like. The term “cycloalkylalkoxycarbonyl” means an acyl groupderived from a cycloalkylalkoxycarboxylic acid of the formulacycloalkylalkyl-O—COOH wherein cycloalkylalkyl has the meaning givenabove. The term “aryloxyalkanoyl” means an acyl radical of the formulaaryl-O-alkanoyl wherein aryl and alkanoyl have the meaning given above.The term “heterocycloalkoxycarbonyl” means an acyl group derived fromheterocycloalkyl-O—COOH wherein heterocycloalkyl is as defined above.The term “heterocycloalkanoyl” is an acyl radical derived from aheterocycloalkylcarboxylic acid wherein heterocyclo has the meaninggiven above. The term “heterocycloalkoxycarbonyl” means an acyl radicalderived from a heterocycloalkyl-O—COOH wherein heterocyclo has themeaning given above. The term “heteroaryloxycarbonyl” means an acylradical derived from a carboxylic acid represented by heteroaryl-O—COOHwherein heteroaryl has the meaning given above. The term “aminocarbonyl”alone or in combination, means an amino-substituted carbonyl (carbamoyl)group wherein the amino group can be a primary, secondary or tertiaryamino group containing substituents selected from alkyl, aryl, aralkyl,cycloalkyl, cycloalkylalkyl radicals and the like. The term“aminoalkanoyl” means an acyl group derived from an amino-substitutedalkylcarboxylic acid wherein the amino group can be a primary, secondaryor tertiary amino group containing substituents selected from alkyl,aryl, aralkyl, cycloalkyl, cycloalkylalkyl radicals and the like. Theterm “halogen” means fluorine, chlorine, bromine or iodine. The term“haloalkyl” means an alkyl radical having the meaning as defined abovewherein one or more hydrogens are replaced with a halogen. Examples ofsuch haloalkyl radicals include chloromethyl, 1-bromoethyl,fluoromethyl, difluoromethyl, trifluoromethyl, 1,1,1-trifluoroethyl andthe like. The term “leaving group” (L or W) generally refers to groupsreadily displaceable by a nucleophile, such as an amine, a thiol or analcohol nucleophile. Such leaving groups are well known in the art.Examples of such leaving groups include, but are not limited to,N-hydroxysuccinimide, N-hydroxybenzotriazole, halides, triflates,tosylates and the like. Preferred leaving groups are indicated hereinwhere appropriate.

Procedures for preparing the compounds of Formula I are set forth below.It should be noted that the general procedure is shown as it relates topreparation of compounds having the specified stereochemistry, forexample, wherein the absolute stereochemistry about the hydroxyl groupis designated as (R). However, such procedures are generally applicableto those compounds of opposite configuration, e.g., where thestereochemistry about the hydroxyl group is (S). In addition, thecompounds having the (R) stereochemistry can be utilized to producethose having the (S) stereochemistry. For example, a compound having the(R) stereochemistry can be inverted to the (S) stereochemistry usingwell-known methods.

Preparation of Compounds of Formula I

The compounds of the present invention represented by Formula I abovecan be prepared utilizing the following general procedures asschematically shown in Schemes I and II.

An N-protected chloroketone derivative of an amino acid having theformula:

wherein P represents an amino protecting group, and R² is as definedabove, is reduced to the corresponding alcohol utilizing an appropriatereducing agent. Suitable amino protecting groups are well known in theart and include carbobenzoxy, t-butoxycarbonyl, and the like. Apreferred amino protecting group is carbobenzoxy. A preferredN-protected chloroketone is N-benzyloxycarbonyl-L-phenylalaninechloromethyl ketone. A preferred reducing agent is sodium borohydride.The reduction reaction is conducted at a temperature of from −10° C. toabout 25° C., preferably at about 0° C., in a suitable solvent systemsuch as, for example, tetrahydrofuran, and the like. The N-protectedchloroketones are commercially available, e.g., such as from Bachem,Inc., Torrance, Calif. Alternatively, the chloroketones can be preparedby the procedure set forth in S. J. Fittkau, J. Prakt. Chem., 315, 1037(1973), and subsequently N-protected utilizing procedures which are wellknown in the art.

The halo alcohol can be utilized directly, as described below, or,preferably, is reacted, preferably at room temperature, with a suitablebase in a suitable solvent system to produce an N-protected aminoepoxide of the formula:

wherein P and R² are as defined above. Suitable solvent systems forpreparing the amino epoxide include ethanol, methanol, isopropanol,tetrahydrofuran, dioxane, and the like including mixtures thereof.Suitable bases for producing the epoxide from the reduced chloroketoneinclude potassium hydroxide, sodium hydroxide, potassium t-butoxide, DBUand the like. A preferred base is potassium hydroxide.

Alternatively, a protected amino epoxide can be prepared, such as inco-owned and co-pending PCT Patent Application Ser. No. PCT/US93/04804(WO 93/23388) and PCT/US94/12201, and each of which is incorporatedherein by reference in their entirety) disclose methods of preparingchiral epoxide, chiral cyanohydrin, chiral amine and other chiralintermediates useful in the preparation of retroviral proteaseinhibitors, starting with a DL-, D- or L-amino acid which is reactedwith a suitable amino-protecting group in a suitable solvent to producean amino-protected amino acid ester. For the purposes of illustration, aprotected L-amino acid with the following formula will be used toprepare the inhibitors of this invention:

wherein P³ represents carboxyl-protecting group, e.g., methyl, ethyl,benzyl, tertiary-butyl, 4-methoxyphenylmethyl and the like; R² is asdefined above; and P¹ and P² and P′ independently are selected fromamine protecting groups, including but not limited to, aralkyl,substituted aralkyl, cycloalkenylalkyl and substitutedcycloalkenylalkyl, allyl, substituted allyl, acyl, alkoxycarbonyl,aralkoxycarbonyl and silyl. Examples of aralkyl include, but are notlimited to benzyl, ortho-methylbenzyl, trityl and benzhydryl, which canbe optionally substituted with halogen, alkyl of C₁–C₈, alkoxy, hydroxy,nitro, alkylene, amino, alkylamino, acylamino and acyl, or their salts,such as phosphonium and ammonium salts. Examples of aryl groups includephenyl, naphthalenyl, indanyl, anthracenyl, durenyl,9-(9-phenylfluorenyl) and phenanthrenyl, cycloalkenylalkyl orsubstituted cycloalkylenylalkyl radicals containing cycloalkyls ofC₆–C₁₀. Suitable acyl groups include carbobenzoxy, t-butoxycarbonyl,iso-butoxycarbonyl, benzoyl, substituted benzoyl, butyryl, acetyl,tri-fluoroacetyl, tri-chloroacetyl, phthaloyl and the like. PreferablyP¹ and P² are independently selected from aralkyl and substitutedaralkyl. More preferably, each of P¹ and P² is benzyl.

Additionally, the P¹ and/or P² and/or P′ protecting groups can form aheterocyclic ring with the nitrogen to which they are attached, forexample, 1,2-bis(methylene)benzene, phthalimidyl, succinimidyl,maleimidyl and the like and where these heterocyclic groups can furtherinclude adjoining aryl and cycloalkyl rings. In addition, theheterocyclic groups can be mono-, di- or tri-substituted, e.g.,nitrophthalimidyl. The term silyl refers to a silicon atom optionallysubstituted by one or more alkyl, aryl and aralkyl groups.

Suitable silyl protecting groups include, but are not limited to,trimethylsilyl, triethylsilyl, tri-isopropylsilyl,tert-butyldimethylsilyl, dimethylphenylsilyl,1,2-bis(dimethylsilyl)benzene, 1,2-bis(dimethylsilyl)ethane anddiphenylmethylsilyl. Silylation of the amine functions to provide mono-or bis-disilylamine can provide derivatives of the aminoalcohol, aminoacid, amino acid esters and amino acid amide. In the case of aminoacids, amino acid esters and amino acid amides, reduction of thecarbonyl function provides the required mono- or bis-silyl aminoalcohol.Silylation of the aminoalcohol can lead to the N,N,O-tri-silylderivative. Removal of the silyl function from the silyl ether functionis readily accomplished by treatment with, for example, a metalhydroxide or ammonium flouride reagent, either as a discrete reactionstep or in situ during the preparation of the amino aldehyde reagent.Suitable silylating agents are, for example, trimethylsilyl chloride,tert-buty-dimethylsilyl chloride, phenyldimethylsilyl chlorie,diphenylmethylsilyl chloride or their combination products withimidazole or DMF. Methods for silylation of amines and removal of silylprotecting groups are well known to those skilled in the art. Methods ofpreparation of these amine derivatives from corresponding amino acids,amino acid amides or amino acid esters are also well known to thoseskilled in the art of organic chemistry including amino acid/amino acidester or aminoalcohol chemistry.

The amino-protected L-amino acid ester is then reduced, to thecorresponding alcohol. For example, the amino-protected L-amino acidester can be reduced with diisobutylaluminum hydride at −78° C. in asuitable solvent such as toluene. Preferred reducing agents includelithium aluminium hydride, lithium borohydride, sodium borohydride,borane, lithium tri-ter-butoxyaluminum hydride, borane/THF complex. Mostpreferably, the reducing agent is diisobutylaluminum hydride (DiBAL-H)in toluene. The resulting alcohol is then converted, for example, by wayof a Swern oxidation, to the corresponding aldehyde of the formula:

wherein P¹, P² and R² are as defined above. Thus, a dichloromethanesolution of the alcohol is added to a cooled (−75 to −68° C.) solutionof oxalyl chloride in dichloromethane and DMSO in dichloromethane andstirred for 35 minutes.

Acceptable oxidizing reagents include, for example, sulfurtrioxide-pyridine complex and DMSO, oxalyl chloride and DMSO, acetylchloride or anhydride and DMSO, trifluoroacetyl chloride or anhydrideand DMSO, methanesulfonyl chloride and DMSO or tetrahydrothiaphene-S-oxide, toluenesulfonyl bromide and DMSO,trifluoromethanesulfonyl anhydride (triflic anhydride) and DMSO,phosphorus pentachloride and DMSO, dimethylphosphoryl chloride and DMSOand isobutyl chloroformate and DMSO. The oxidation conditions reportedby Reetz et al [Agnew Chem., 99, p. 1186, (1987)], Agnew Chem. Int. Ed.Engl., 26, p. 1141, 1987) employed oxalyl chloride and DMSO at −78° C.

The preferred oxidation method described in this invention is sulfurtrioxide pyridine complex, triethylamine and DMSO at room temperature.This system provides excellent yields of the desired chiral protectedamino aldehyde usable without the need for purification i.e., the needto purify kilograms of intermediates by chromatography is eliminated andlarge scale operations are made less hazardous. Reaction at roomtemperature also eliminated the need for the use of low temperaturereactor which makes the process more suitable for commercial production.

The reaction may be carried out under an inert atmosphere such asnitrogen or argon, or normal or dry air, under atmospheric pressure orin a sealed reaction vessel under positive pressure. Preferred is anitrogen atmosphere. Alternative amine bases include, for example,tri-butyl amine, tri-isopropyl amine, N-methylpiperidine, N-methylmorpholine, azabicyclononane, diisopropylethylamine,2,2,6,6-tetramethylpiperidine, N,N-dimethylaminopyridine, or mixtures ofthese bases. Triethylamine is a preferred base. Alternatives to pureDMSO as solvent include mixtures of DMSO with non-protic or halogenatedsolvents such as tetrahydrofuran, ethyl acetate, toluene, xylene,dichloromethane, ethylene dichloride and the like. Dipolar aproticco-solvents include acetonitrile, dimethylformamide, dimethylacetamide,acetamide, tetramethyl urea and its cyclic analog, N-methylpyrrolidone,sulfolane and the like. Rather than N,N-dibenzylphenylalaninol as thealdehyde precursor, the phenylalaninol derivatives discussed above canbe used to provide the corresponding N-monosubstituted [either P¹ orP²=H] or N,N-disubstituted aldehyde.

In addition, hydride reduction of an amide or ester derivative of thecorresponding benzyl (or other suitable protecting group) nitrogenprotected phenylalanine, substituted phenylalanine or cycloalkyl analogof phenylalanine derivative can be carried out to provide the aldehydes.Hydride transfer is an additional method of aldehyde synthesis underconditions where aldehyde condensations are avoided, cf, OppenauerOxidation.

The aldehydes of this process can also be prepared by methods ofreducing protected phenylalanine and phenylalanine analogs or theiramide or ester derivatives by, e.g., sodium amalgam with HCl in ethanolor lithium or sodium or potassium or calcium in ammonia. The reactiontemperature may be from about −20° C. to about 45° C., and preferablyfrom abut 5° C. to about 25° C. Two additional methods of obtaining thenitrogen protected aldehyde include oxidation of the correspondingalcohol with bleach in the presence of a catalytic amount of2,2,6,6-tetramethyl-1-pyridyloxy free radical. In a second method,oxidation of the alcohol to the aldehyde is accomplished by a catalyticamount of tetrapropylammonium perruthenate in the presence ofN-methylmorpholine-N-oxide.

Alternatively, an acid chloride derivative of a protected phenylalanineor phenylalanine derivative as disclosed above can be reduced withhydrogen and a catalyst such as Pd on barium carbonate or bariumsulphate, with or without an additional catalyst moderating agent suchas sulfur or a thiol (Rosenmund Reduction).

The aldehyde resulting from the Swern oxidation is then reacted with ahalomethyllithium reagent, which reagent is generated in situ byreacting an alkyllithium or arylithium compound with a dihalomethanerepresented by the formula X¹CH₂X² wherein X¹ and X² independentlyrepresent I, Br or Cl. For example, a solution of the aldehyde andchloroiodomethane in THF is cooled to −78° C. and a solution ofn-butyllithium in hexane is added. The resulting product is a mixture ofdiastereomers of the corresponding amino-protected epoxides of theformulas:

The diastereomers can be separated e.g., by chromatography, or,alternatively, once reacted in subsequent steps the diastereomericproducts can be separated. A D-amino acid can be utilized in place ofthe L-amino acid in order to prepare compounds having an (S)stereochemistry at the carbon bonded to R².

The addition of chloromethylithium or bromomethylithium to a chiralamino aldehyde is highly diastereoselective. Preferably, thechloromethyllithium or bromomethylithium is generated in-situ from thereaction of the dihalomethane and n-butyllithium. Acceptablemethyleneating halomethanes include chloroiodomethane,bromochloromethane, dibromomethane, diiodomethane, bromofluoromethaneand the like. The sulfonate ester of the addition product of, forexample, hydrogen bromide to formaldehyde is also a methyleneatingagent. Tetrahydrofuran is the preferred solvent, however alternativesolvents such as toluene, dimethoxyethane, ethylene dichloride,methylene chloride can be used as pure solvents or as a mixture. Dipolaraprotic solvents such as acetonitrile, DMF, N-methylpyrrolidone areuseful as solvents or as part of a solvent mixture. The reaction can becarried out under an inert atmosphere such as nitrogen or argon. Forn-butyl lithium can be substituted other organometalic reagents reagentssuch as methyllithium, tert-butyl lithium, sec-butyl lithium,phenyllithium, phenyl sodium and the like. The reaction can be carriedout at temperatures of between about −80° C. to 0° C. but preferablybetween about −80° C. to −20° C. The most preferred reactiontemperatures are between −40° C. to −15° C. Reagents can be added singlybut multiple additions are preferred in certain conditions. Thepreferred pressure of the reaction is atmospheric however a positivepressure is valuable under certain conditions such as a high humidityenvironment.

Alternative methods of conversion to the epoxides of this inventioninclude substitution of other charged methylenation precurser speciesfollowed by their treatment with base to form the analogous anion.Examples of these species include trimethylsulfoxonium tosylate ortriflate, tetramethylammonium halide, methyldiphenylsulfoxonium halidewherein halide is chloride, bromide or iodide.

The conversion of the aldehydes of this invention into their epoxidederivative can also be carried out in multiple steps. For example, theaddition of the anion of thioanisole prepared from, for example, a butylor aryl lithium reagent, to the protected aminoaldehyde, oxidation ofthe resulting protected aminosulfide alcohol with well known oxidizingagents such as hydrogen peroxide, tert-butyl hypochlorite, bleach orsodium periodate to give a sulfoxide. Alkylation of the sulfoxide with,for example, methyl iodide or bromide, methyl tosylate, methyl mesylate,methyl triflate, ethyl bromide, isopropyl bromide, benzyl chloride orthe like, in the presence of an organic or inorganic base Alternatively,the protected aminosulfide alcohol can be alkylated with, for example,the alkylating agents above, to provide a sulfonium salts that aresubsequently converted into the subject epoxides with tert-amine ormineral bases.

The desired epoxides formed, using most preferred conditions,diastereoselectively in ratio amounts of at least about an 85:15 ratio(S:R). The product can be purified by chromatography to give thediastereomerically and enantiomerically pure product but it is moreconveniently used directly without purification to prepare retroviralprotease inhibitors. The foregoing process is applicable to mixtures ofoptical isomers as well as resolved compounds. If a particular opticalisomer is desired, it can be selected by the choice of startingmaterial, e.g., L-phenylalanine, D-phenylalanine, L-phenylalaninol,D-phenylalaninol, D-hexahydrophenylalaninol and the like, or resolutioncan occur at intermediate or final steps. Chiral auxiliaries such as oneor two equivilants of camphor sulfonic acid, citric acid, camphoricacid, 2-methoxyphenylacetic acid and the like can be used to form salts,esters or amides of the compounds of this invention. These compounds orderivatives can be crystallized or separated chromatographically usingeither a chiral or achiral column as is well known to those skilled inthe art.

The amino epoxide is then reacted, in a suitable solvent system, with anequal amount, or preferably an excess of, a desired amine of the formulaR³NH₂, wherein R³ is hydrogen or is as defined above. The reaction canbe conducted over a wide range of temperatures, e.g., from about 10° C.to about 100° C., but is preferably, but not necessarily, conducted at atemperature at which the solvent begins to reflux. Suitable solventsystems include protic, non-protic and dipolar aprotic organic solventssuch as, for example, those wherein the solvent is an alcohol, such asmethanol, ethanol, isopropanol, and the like, ethers such astetrahydrofuran, dioxane and the like, and toluene,N,N-dimethylformamide, dimethyl sulfoxide, and mixtures thereof. Apreferred solvent is isopropanol. The resulting product is a3-(N-protected amino)-3-(R²)-1-(NHR³)-propan-2-ol derivative(hereinafter referred to as an amino alcohol) can be represented by theformulas:

wherein P, P¹, P², R² and R³ are as described above. Alternatively, ahaloalcohol can be utilized in place of the amino epoxide.

The amino alcohol defined above is then reacted in a suitable solventwith the sulfonyl chloride R⁴SO₂Cl, the sulfonyl bromide R⁴SO₂Br or thecorresponding sulfonyl anhydride, preferably in the presence of an acidscavenger. Suitable solvents in which the reaction can be conductedinclude methylene chloride, tetrahydrofuran and the like. Suitable acidscavengers include triethylamine, pyridine and the like. The resultingsulfonamide derivative can be represented, depending on the epoxideutilized by the formulas

wherein P, P¹, P², R², R³ and R⁴ are as defined above. Theseintermediates are useful for preparing inhibitor compounds of thepresent invention.

Alternatively, the protected amino alcohol from the epoxide opening canbe further protected at the newly introduced amino group with aprotecting group P′ which is not removed with the removal of the aminoprotecting groups P or P¹ and P², i.e., P′ is selectively removable. Oneskilled in the art can choose appropriate combinations of P′, P, P¹ andP². For example, suitable combinations are P=Cbz and P′=Boc; P′=Cbz andP=Boc; P¹=Cbz, P²=benzyl and P′=Boc; and P¹=P²=benzyl and P′=Boc. Theresulting compound represented by the formula

can be carried through the remainder of the synthesis to provide acompound of the formula

wherein n, t, P′, R¹, R², R³ and R⁵ are as defined above. The remainderof the synthesis above can be carried out as desired either by theaddition of desired residues or groups one at a time or in a preformedmolecule made up of more that one residue or group in one step. Theformer approach is the sequential synthesis method and the latter is theconvergent synthesis method. Synthetic transformations are possible atthis stage also. The protecting group P′ is then selectively removed andthe resulting amine is reacted with the sulfonyl chloride R⁴SO₂Cl, thesulfonyl bromide R⁴SO₂Br or the corresponding sulfonyl anhydride,preferably in the presence of an acid scavenger, to form the compoundsof the present invention

wherein n, t, R¹, R², R³, R⁴ and R⁵ are as defined above. This selectivedeprotection and conversion to the sulfonamide can be accomplished ateither the end of the synthesis or at any appropriate intermediate stepas desired. An example is outlined in Scheme II.

The sulfonyl halides of the formula R⁴SO₂X can be prepared by thereaction of a suitable aryl, heteroaryl and benzo fused heterocycloGrignard or lithium reagents with sulfuryl chloride, or sulfur dioxidefollowed by oxidation with a halogen, preferably chlorine. Aryl,heteroaryl and benzo fused heterocyclo Grignard or lithium reagents canbe prepared from their corresponding halide (such as chloro or bromo)compounds which are commercially available or readily prepared fromcommercially available starting materials using known methods in theart. Also, thiols may be oxidized to sulfonyl chlorides using chlorinein the presence of water under carefully controlled conditions.Additionally, sulfonic acids, such as arylsulfonic acids, may beconverted to sulfonyl halides using reagents such as PCl₅, SOCl₂,ClC(O)C(O)Cl and the like, and also to anhydrides using suitabledehydrating reagents. The sulfonic acids may in turn be prepared usingprocedures well known in the art. Some sulfonic acids are commerciallyavailable. In place of the sulfonyl halides, sulfinyl halides (R⁴SOX) orsulfenyl halides (R⁴SX) can be utilized to prepare compounds wherein the—SO₂— moiety is replaced by an —SO— or —S— moiety, respectively.Arylsulfonic acids, benzo fused heterocyclo sulfonic acids or heteroarylsulfonic acids can be prepared by sulfonation of the aromatic ring bywell known methods in the art, such as by reaction with sulfuric acid,SO₃, SO₃ complexes, such as DMF(SO₃), pyridine(SO₃),N,N-dimethylacetamide(SO₃), and the like. Preferably, arylsulfonylhalides are prepared from aromatic compounds by reaction with DMF(SO₃)and SOCl₂ or ClC(O)C(O)Cl. The reactions may be performed stepwise or ina single pot.

Arylsulfonic acids, benzo fused heterocyclo sulfonic acids, heteroarylsulfonic acids, arylmercaptans, benzo fused heterocyclo mercaptans,heteroarylmercaptans, arylhalides, benzo fused heterocyclo halides,heteroarylhalides, and the like are commercially available or can bereadily prepared from starting materials commercially available usingstandard methods well known in the art. For example, a number ofsulfonic acids (R⁴SO₃H) represented by the formulas

wherein A, B, Z, R⁶, R⁷ and R⁹ are as defined above, have been preparedfrom 1,2-benzenedithiol, 2-mercaptanphenol, 1,2-benzenediol,2-aminobenzothiazole, benzothiazole, 2-aminobenzimidazole,benzimidazole, and the like, which are commercially available, byCarter, U.S. Pat. No. 4,595,407; Ehrenfreund et al., U.S. Pat. No.4,634,465; Yoder et al., J. Heterocycl. Chem. 4:166–167 (1967); Cole etal., Aust. J. Chem. 33:675–680 (1980); Cabiddu et al., Synthesis 797–798(1976); Ncube et al., Tet. Letters 2345–2348 (1978); Ncube et al., Tet.Letters 255–256 (1977); Ansink & Cerfontain, Rec. Trav. Chim.Pays-Bas108:395–403 (1989); and Kajihara & Tsuchiya, EP 638564 A1, each of whichare incorporated herein by reference in their entirety. For example,1,2-benzenedithiol, 2-mercaptanphenol or 1,2-benzenediol can be reactedwith R⁶R⁷C(L′)₂, where L′ is as defined below, preferably, Br or I, inthe presence of a base, such as hydroxide, or R⁶R⁷C═O in the presence ofacid, such as toluenesulfonic acid, or P₂O₅., to prepare the substitutedbenzo fused heterocycle of formula

which can then be sulfonylated to the sulfonic acid above. For example,CF₂Br₂ or CD₂Br₂ can be reacted with 1,2-benzenedithiol,2-mercaptanphenol or 1,2-benzenediol in the presence of base to producethe compounds

respectively, wherein A and B are O or S and D is a deuterium atom.Also, when A and/or B represent S, the sulfur can be oxidized using themethods described below to the sulfone or sulfoxide derivatives.

Following preparation of the sulfonamide derivative, the aminoprotecting group P or P¹ and P² amino protecting groups are removedunder conditions which will not affect the remaining portion of themolecule. These methods are well known in the art and include acidhydrolysis, hydrogenolysis and the like. A preferred method involvesremoval of the protecting group, e.g., removal of a carbobenzoxy group,by hydrogenolysis utilizing palladium on carbon in a suitable solventsystem such as an alcohol, acetic acid, and the like or mixturesthereof. Where the protecting group is a t-butoxycarbonyl group, it canbe removed utilizing an inorganic or organic acid, e.g., HCl ortrifluoroacetic acid, in a suitable solvent system, e.g., dioxane ormethylene chloride. The resulting product is the amine salt derivative.

Following neutralization of the salt, the amine is then coupled to thesulfone/sulfoxidealkanoyl compound or an optical isomer thereof (such aswhere the group —CH(R¹)— is R or S), corresponding to the formula

wherein n, t and R⁵ are as defined above, and L is leaving group such ashalide, anhydride, active ester, and the like. Alternatively, thesulfone/sulfoxide alkanoyl compound or an optical isomer thereof can becoupled to the protected amine

followed by deprotection and coupling to R⁴SO₂X, where X is Cl or Br andP′, R², R³ and R⁴ is as defined above.

Such sulfone/sulfoxidealkanoyl compounds where n is 1 can be prepared byreacting a mercaptan of the formula R⁵SH with a substituted methacrylateof the formula

by way of a Michael Addition. Such substituted methacrylates arecommercially available or readily prepared from commercially availablestarting materials using standard methods well known in the art. TheMichael Addition is conducted in a suitable solvent and in the presenceof a suitable base, to produce the corresponding thiol derivativerepresented by the formula

wherein P³, R¹ and R⁵ are as defined above. Suitable solvents in whichthe Michael Addition can be conducted include alcohols such as, forexample, methanol, ethanol, butanol and the Like, as well as ethers,e.g., THF, and acetonitrile, DMF, DMSO, and the like, including mixturesthereof. Suitable bases include Group I metal alkoxides such as, forexample sodium methoxide, sodium ethoxhide, sodium butoxide and the likeas well as Group I metal hydrides, such as sodium hydride, includingmixtures thereof. The thiol derivative is converted into thecorresponding sulfone or sulfoxide of the formula

by oxidizing the thiol derivative with a suitable oxidation agent in asuitable solvent. Suitable oxidation agents include, for example,hydrogen peroxide, sodium meta-perborate, oxone (potassium peroxymonosulfate), meta-chloroperoxybenzoic acid, periodic acid and the like,including mixtures thereof. Suitable solvents include acetic acid (forsodium meta-perborate) and, for other peracids, ethers such as THF anddioxane, and acetonitrile, DMF and the like, including mixtures thereof.

The sulfone/sulfoxide is then converted into the corresponding free acidof the formula

utilizing a suitable base, e.g., lithium hydroxide, sodium hydroxide,and the like, including mixtures thereof, in a suitable solvent, suchas, for example, THF, acetonitrile, DMF, DMSO, methylene chloride andthe like, including mixtures thereof. The free acid can then beconverted into the sulfone/sulfoxidealkanoyl compound

wherein n, t and R⁵ are as defined above, and L is leaving group such ashalide, anhydride, active ester, and the like. Alternatively, the freeacid can be resolved into its optical isomers (such as where the group—CH(R¹)— is R or S) using well known methods in the art, such as byforming diastereomeric salts or esters and crystallizing orchromatographing, and then converted into the sulfone/sulfoxidealkanoylcompound.

Alternatively, the thioether or corresponding protected thiol offormulas

respectively, where n, L, R¹ and R⁵ are as defined above, can be coupledto one of the amines

followed by conversion to the protease inhibitors of the presentinvention. P⁴ is a sulfur protecting group, such as acetyl, benzoyl andthe like. The acetyl and benzoyl groups can be removed by treatment withan inorganic base or an amine, preferably ammonia, in an appropriatesolvent such as methanol, ethanol, isopropanol, toluene ortetrahydrofuran. The preferred solvent is methanol.

For example, one can couple the commercially available acid

to one of the amines

deacetylate the sulfur group, such as by hydrolysis with a suitablebase, such as hydroxide, or an amine, such as ammonia, and then reactthe resulting thiol with R⁵L′ agent, wherein R⁵ and L′ are as definedabove, to afford compounds with one of the following structures

or specific diastereomeric isomers thereof. The sulfur can then beoxidized to the corresponding sulfone or sulfoxide using suitableoxidizing agents, as described above, to afford the desired intermediatefollowed by further reactions to prepare the sulfonamide inhibitor, ordirectly to the sulfonamide inhibitor. Alternatively, the acid or the P³protected acid can be deacetylated, reacted with R⁵L′ agent, deprotectedand oxidized to the corresponding sulfone or sulfoxide using suitableoxidizing agents, as described above to afford the compound of formula

wherein t and R⁵ are as defined above. This sulfone/sulfoxide carboxylicacid can then be coupled to the amine intermediate described abovefollowed by further reaction to prepare the sulfonamide inhibitor, or tothe sulfonamide amine compound to produce the sulfonamide inhibitordirectly. The L′ group of the R⁵L′ agent is a leaving group such as ahalide (chloride, bromide, iodide), mesylate, tosylate or triflate. Thereaction of the mercaptan with R⁵L′ is performed in the presence of asuitable base, such as triethylamine, diisopropylethylamine,1,8-diazabicyclo[5.4.0] undec-7-ene (DBu) and the like, in a suitablesolvent such as toluene, tetrahydrofuran, or methylene chloride. Thepreferred base is DBU and the preferred solvent is toluene. Where R⁵ isa methyl group, R⁵L′ can be methyl chloride, methyl bromide, methyliodide, or dimethyl sulfate, and preferably methyl iodide.

Alternatively, a substituted methacrylate of the formula

wherein L′ represents a leaving group as previously defined, P³ is asdefined above and R¹⁰ represents radicals which upon reduction of thedouble bond produce radicals of R¹, is reacted with R⁵SM followed byoxidation, as described above, or a suitable sulfonating agent, such as,for example, a sulfinic acid represented by the formula R⁵SO₂M, whereinR⁵ is as defined above and M represents a metal adapted to form a saltof the acid, e.g., sodium, to produce the corresponding sulfonerepresented by the formula

wherein P³, R⁵ and R¹⁰ are as defined above. The sulfone is thendeprotected to form the corresponding carboxylic acid. For example, whenP³ is a tertiary-butyl group, it can be removed by treatment with anacid, such as hydrochloric acid or trifluoracetic acid. The preferredmethod is using 4N hydrochloric acid in dioxane.

The resulting carboxylic acid is then asymmetrically hydrogenatedutilizing an asymmetric hydrogenation catalyst such as, for example, aruthenium-BINAP complex, to produce the reduced product, substantiallyenriched in the more desired isomer, represented by one of the formulas

wherein R¹ and R⁵ are as defined above. Where the more active isomer hasthe R-stereochemistry, a Ru(R-BINAP) asymmetric hydrogenation catalystcan be utilized. Conversely, where the more active isomer has theS-sterochemistry, a Ru(S-BINAP) catalyst can be utilized. Where bothisomers are active, or where it is desired to have a mixture of the twodiastereomers, a hydrogenation catalyst such as platinum or palladium oncarbon can be utilized to reduce the above compound. The reducedcompound is then coupled to an amine as described above.

The chemical reactions described above are generally disclosed in termsof their broadest application to the preparation of the compounds ofthis invention. Occasionally, the reactions may not be applicable asdescribed to each compound included within the disclosed scope. Thecompounds for which this occurs will be readily recognized by thoseskilled in the art. In all such cases, either the reactions can besuccessfully performed by conventional modifications known to thoseskilled in the art, e.g., by appropriate protection of interferinggroups, by changing to alternative conventional reagents, by routinemodification of reaction conditions, and the like, or other reactionsdisclosed herein or otherwise conventional, will be applicable to thepreparation of the corresponding compounds of this invention. In allpreparative methods, all starting materials are known or readilyprepared from known starting materials.

Without further elaboration, it is believed that one skilled in the artcan, using the preceding description, utilize the present invention toits fullest extent. The following preferred specific embodiments are,therefore, to be construed as merely illustrative, and not limitative ofthe remainder of the disclosure in any way whatsoever.

All reagents were used as received without purification. All proton andcarbon NMR spectra were obtained on either a Varian VXR-300 or VXR-400nuclear magnetic resonance spectrometer.

The following Examples illustrate the preparation of inhibitor compoundsof the present invention and intermediates useful in preparing theinhibitor compounds of the present invention.

EXAMPLE 1

Preparation of 2S-[Bis(phenylmethyl)amino]benzenepropanol

Method 1: 2S-[Bis(phenylmethyl)amino]benzenepropanol from the DIBALReduction of N,N-bis(phenylmethyl)-L-Phenylalanine phenylmethyl ester

Step 1:

A solution of L-phenylalanine (50.0 g, 0.302 mol) sodium hydroxide (24.2g, 0.605 mol) and potassium carbonate (83.6 g, 0.605 mol) in water (500mL) was heated to 97° C. Benzyl bromide (108.5 mL, 0.605 then slowlyadded (addition time—25 min). The mixture was stirred at 97° C. for 30minutes under a nitrogen atmosphere. The solution was cooled to roomtemperature and extracted with toluene (2×250 mL). The combined organiclayers were washed with water and brine, dried over magnesium sulfate,filtered and concentrated to an oil. The identity of the product wasconfirmed as follows. Analytical TLC (10% ethyl acetate/hexane, silicagel) showed major component at Rf value=0.32 to be the desiredtribenzylated compound, N,N-bis(phenylmethyl)-L-phenylalaninephenylmethyl ester. This compound can be purified by columnchromatography (silica gel, 15% ethyl acetate/hexane). Usually theproduct is pure enough to be used directly in the next step withoutfurther purification. ¹H NMR spectrum was in agreement with publishedliterature. ¹H NMR (CDCL₃) ∂, 3.00 and 3.14 (ABX-system, 2H, J_(AB)=14.1Hz, J_(AX)=7.3 Hz and J_(BX)=5.9 Hz), 3.54 and 3.92 (AB-System, 4H,J_(AB)=13.9 Hz), 3.71 (t, 1H, J=7.6 Hz), 5.11 and 5.23 (AB-System, 2H,J_(AB)=12.3 Hz), and 7.18 (m, 20H). EIMS: m/z 434 (M−1).

Step 2:

The benzylated phenylalanine phenylmethyl ester (0.302 mol) from theprevious reaction was dissolved in toluene (750 mL) and cooled to −55°C. A 1.5 M solution of DIBAL in toluene (443.9 mL, 0 666 mol) was addedat a rate to maintain the temperature between −55 to −50° C. (additiontime—1 hr). The mixture was stirred for 20 minutes under a nitrogenatmosphere and then quenched at −55° C. by the slow addition of methanol(37 ml). The cold solution was then poured into cold (5° C.) 1.5 N HClsolution (1.8 L). The precipitated solid (approx. 138 g) was filteredoff and washed with toluene. The solid material was suspended in amixture of toluene (400 mL) and water (100 ml). The mixture was cooledto 5° C. and treated with 2.5 N NaOH (186 mL) and then stirred at roomtemperature until solid dissolved. The toluene layer was separated fromthe aqueous phase and washed with water and brine, dried over magnesiumsulfate, filtered and concentrated to a volume of 75 mL (89 g). Ethylacetate (25 mL) and hexane (25 mL) were added to the residue upon whichthe desired alcohol product began to crystallize. After 30 min, anadditional 50 mL hexane were added to promote further crystallization.The solid was filtered off and washed with 50 mL hexane to give 34.9 gof first crop product. A second crop of product (5.6 g) was isolated byrefiltering the mother liquor. The two crops were combined andrecrystallized from ethyl acetate (20 mL) and hexane (30 mL) to give 40g of βS-2-[Bis(phenyl-methyl)amino]benzenepropanol, 40% yield fromL-phenylalanine. An additional 7 g (7%) of product can be obtained fromrecrystallization of the concentrated mother liquor. TLC of productRf=0.23 (10% ethyl acetate/hexane, silica gel); ¹H NMR (CDCl₃) ∂ 2.44(m, 1H,), 3.09 (m, 2H), 3.33 (m, 1H), 3.48 and 3.92 (AB-System, 4H,J_(AB)=13.3 Hz), 3.52 (m, 1H) and 7.23 (m, 15H); [α]_(D)25+42.4 (c 1.45,CH₂Cl₂); DSC 77.67° C.; Anal. Calcd. for C₂₃H₂₅ON: C, 83.34; H, 7.60; N,4.23. Found: C, 83.43; H, 7.59; N, 4.22. HPLC on chiral stationaryphase: Cyclobond I SP column (250×4.6 mm I.D.), mobile phase:methanol/triethyl ammonium acetate buffer pH 4.2 (58:42, v/v), flow-rateof 0.5 ml/min, detection with detector at 230 nm and a temperature of 0°C. Retention time: 11.25 min., retention time of the desired productenantiomer: 12.5 min.

Method 2: Preparation of βS-2-[Bis(phenylmethyl)amino]benzene-propanolfrom the N,N-Dibenzylation of L-Phenylalaninol

L-phenylalaninol (176.6 g, 1.168 mol) was added to a stirred solution ofpotassium carbonate (484.6 g, 3.506 mol) in 710 mL of water. The mixturewas heated to 65° C. under a nitrogen atmosphere. A solution of benzylbromide (400 g, 2.339 mol) in 3A ethanol (305 mL) was added at a ratethat maintained the temperature between 60–68° C. The biphasic solutionwas stirred at 65° C. for 55 min and then allowed to cool to 10° C. withvigorous stirring. The oily product solidified into small granules. Theproduct was diluted with 2.0 L of tap water and stirred for 5 minutes todissolve the inorganic by products. The product was isolated byfiltration under reduced pressure and washed with water until the pH is7. The crude product obtained was air dried overnight to give a semi-drysolid (407 g) which was recrystallized from 1.1 L of ethylacetate/heptane (1:10 by volume). The product was isolated by filtration(at −8° C.), washed with 1.6 L of cold (−10° C.) ethyl acetate/heptane(1:10 by volume) and air-dried to give 339 g (88% yield) ofβS-2-[Bis(phenylmethyl)amino]benzene-propanol, Mp=71.5–73.0° C. Moreproduct can be obtained from the mother liquor if necessary. The otheranalytical characterization was identical to compound prepared asdescribed in Method 1.

EXAMPLE 2

Preparation of 2S-[Bis(phenylmethyl)amino]benzeneproyanaldehyde

Method 1:

2S-[Bis(phenylmethyl)amino]benzene-propanol (200 g, 0.604 mol) wasdissolved in triethylamine (300 mL, 2.15 mol). The mixture was cooled to12° C. and a solution of sulfur trioxide/pyridine complex (380 g, 2.39mol) in DMSO (1.6 L) was added at a rate to maintain the temperaturebetween 8–17° C. (addition time—1.0 h). The solution was stirred atambient temperature under a nitrogen atmosphere for 1.5 hour at whichtime the reaction was complete by TLC analysis (33% ethylacetate/hexane, silica gel). The reaction mixture was cooled with icewater and quenched with 1.6 L of cold water (10–15° C.) over 45 minutes.The resultant solution was extracted with ethyl acetate (2.0 L), washedwith 5% citric acid (2.0 L), and brine (2.2 L), dried over MgSO₄ (280 g)and filtered. The solvent was removed on a rotary evaporator at 35–40°C. and then dried under vacuum to give 198.8 g of2S-[Bis-(phenylmethyl)amino]-benzenepropanaldehyde as a pale yellow oil(99.9%). The crude product obtained was pure enough to be used directlyin the next step without purification. The analytical data of thecompound were consistent with the published literature.[α]_(D)25=−92.9°(c 1.87, CH₂Cl₂); ¹H NMR (400 MHz, CDCl₃) ∂, 2.94 and 3.15 (ABX-System,2H, J_(AB)=13.9 Hz, J_(AX)=7.3 Hz and J_(BX)=6.2 Hz), 3.56 (t, 1H, 7.1Hz), 3.69 and 3.82 (AB-System, 4H, J_(AB)=13.7 Hz), 7.25 (m, 15 H) and9.72 (s, 1H); HRMS Calcd for (M+1) C₂₃H₂₄NO 330.450, found: 330.1836.Anal. Calcd. for C₂₃H₂₃ON: C, 83.86; H, 7.04; N, 4.25. Found: C, 83.64;H, 7.42; N, 4.19. HPLC on chiral stationary phase: (S,S) Pirkle-Whelk-O1 column (250×4.6 mm I.D.), mobile phase: hexane/isopropanol (99.5:0.5,v/v), flow-rate: 1.5 ml/min, detection with UV detector at 210 nm.Retention time of the desired S-isomer: 8.75 min., retention time of theR-enantiomer 10.62 min.

Method 2:

A solution of oxalyl chloride (8.4 ml, 0.096 mol) in dichloromethane(240 ml) was cooled to −74° C. A solution of DMSO (12.0 ml, 0.155 mol)in dichloromethane (50 ml) as then slowly added at a rate to maintainthe temperature at −74° C. (addition time—1.25 hr). The mixture wasstirred for 5 min. followed by addition of a solution ofβS-2-[bis(phenylmethyl)amino]benzene-propanol (0.074 mol) in 100 ml ofdichloromethane (addition time −20 min., temp. −75° C. to −68° C). Thesolution was stirred at −78° C. for 35 minutes under a nitrogenatmosphere. Triethylamine (41.2 ml, 0.295 mol) was then added over 10min. (temp. −78° to −68° C.) upon which the ammonium salt precipitated.The cold mixture was stirred for 30 min. and then water (225 ml) wasadded. The dichloromethane layer was separated from the aqueous phaseand washed with water, brine, dried over magnesium sulfate, filtered andconcentrated. The residue was diluted with ethyl acetate and hexane andthen filtered to further remove the ammonium salt. The filtrate wasconcentrated to give αS-[bis(phenylmethyl)amino]benzenepropanaldehyde.The aldehyde was carried on to the next step without purification.

Method 3:

To a mixture of 1.0 g(3.0 mmoles) ofβS-2-[bis(phenylmethyl)amino]benzenepropanol 0.531 g(4.53 mmoles) ofN-methyl morpholine, 2.27 g of molecular sieves(4A) and 9.1 mL ofacetonitrile was added 53 mg (0.15 mmoles) of tetrapropylammoniumperruthenate (TPAP). The mixture was stirred for 40 minutes at roomtemperature and concentrated under reduced pressure. The residue wassuspended in 15 mL of ethyl acetate, filtered through a pad of silicagel. The filtrate was concentrated under reduced pressure to give aproduct containing approximately 50% ofαS-2-[bis(phenylmethyl)amino]benzene propanaldehyde as a pale yellowoil.

Method 4:

To a solution of 1.0 g (3.02 mmoles) ofβS-2-[bis(phenylmethyl)amino]benzenepropanol in 9.0 mL of toluene wasadded 4.69 mg(0.03 mmoles) of 2,2,6,6-tetramethyl-1-piperidinyloxy, freeradical (TEMPO), 0.32 g(3.11 mmoles) of sodium bromide, 9.0 mL of ethylacetate and 1.5 mL of water. The mixture was cooled to 0° C. and anaqueous solution of 2.87 mL of 5% household bleach containing 0.735g(8.75 mmoles) of sodium bicarbonate and 8.53 mL of water was addedslowly over 25 minutes. The mixture was stirred at 0° C. for 60 minutes.Two more additions (1.44 mL each) of bleach was added followed bystirring for 10 minutes. The two phase mixture was allowed to separate.The aqueous layer was extracted twice with 20 mL of ethyl acetate. Thecombined organic layer was washed with 4.0 mL of a solution containing25 mg of potassium iodide and water(4.0 mL), 20 mL of 10% aqueous sodiumthiosulfate solution and then brine solution. The organic solution wasdried over magnesium sulfate, filtered and concentrated under reducedpressure to give 1.34 g of crude oil containing a small amount of thedesired product aldehyde,αS-[bis(phenylmethyl)amino]benzenepropanaldehyde.

Method 5:

Following the same procedures as described in Method 1 of this Exampleexcept 3.0 equivalents of sulfur trioxide pyridine complex was used andαS-[bis(phenylmethyl)amino]benzenepropanaldehyde was isolated incomparable yields.

EXAMPLE 3

Preparation of N,N-dibenzyl-3(S)-amino-1,2-(S)-epoxy-4-phenylbutane

Method 1:

A solution of αS-[Bis(phenylmethyl)amino]benzene-propanaldehyde (191.7g, 0.58 mol) and chloroiodomethane (56.4 mL, 0.77 mol) intetrahydrofuran (1.8 L) was cooled to −30 to −35° C. (colder temperaturesuch as −70° C. also worked well but warmer temperatures are morereadily achieved in large scale operations) in a stainless steel reactorunder a nitrogen atmosphere. A solution of n-butyl lithium in hexane(1.6 M, 365 mL, 0.58 mol) was then added at a rate that maintained thetemperature below −25° C. After addition the mixture was stirred at −30to −35° C. for 10 minutes. More additions of reagents were carried outin the following manner: (1) additional chloroiodomethane (17 mL) wasadded, followed by n-butyl lithium (110 mL) at <−25° C. After additionthe mixture was stirred at −30 to −35° C. for 10 minutes. This wasrepeated once. (2) Additional chloroiodomethane (8.5 mL, 0.11 mol) wasadded, followed by n-butyl lithium (55 mL, 0.088 mol) at <−25° C. Afteraddition the mixture was stirred at −30 to −35° C. for 10 minutes. Thiswas repeated 5 times. (3) Additional chloroiodomethane (8.5 mL, 0.11mol) was added, followed by n-butyl lithium (37 mL, 0.059 mol) at <−25°C. After addition the mixture was stirred at −30 to −35° C. for 10minutes. This was repeated once. The external cooling was stopped andthe mixture warmed to ambient temp. over 4 to 16 hours when TLC (silicagel, 20% ethyl acetate/hexane) indicated that the reaction wascompleted. The reaction mixture was cooled to 10° C. and quenched with1452 g of 16% ammonium chloride solution (prepared by dissolving 232 gof ammonium chloride in 1220 mL of water), keeping the temperature below23° C. The mixture was stirred for 10 minutes and the organic andaqueous layers were separated. The aqueous phase was extracted withethyl acetate (2×500 mL). The ethyl acetate layer was combined with thetetrahydrofuran layer. The combined solution was dried over magnesiumsulfate (220 g), filtered and concentrated on a rotary evaporator at 65°C. The brown oil residue was dried at 70° C. in vacuo (0.8 bar) for 1 hto give 222.8 g of crude material. (The crude product weight was >100%.Due to the relative instability of the product on silica gel, the crudeproduct is usually used directly in the next step without purification).The diastereomeric ratio of the crude mixture was determined by protonNMR: (2S)/(2R): 86:14. The minor and major epoxide diastereomers werecharacterized in this mixture by tlc analysis (silica gel, 10% ethylacetate/hexane), Rf=0.29 & 0.32, respectively. An analytical sample ofeach of the diastereomers was obtained by purification on silica-gelchromatography (3% ethyl acetate/hexane) and characterized as follows:

N,N,αS-Tris(phenylmethyl)-2S-oxiranemethanamine

¹H NMR (400 MHz, CDCl₃) ∂ 2.49 and 2.51 (AB-System, 1H, J_(AB)=2.82),2.76 and 2.77 (AB-System, 1H, J_(AB)=4.03), 2.83 (m, 2H), 2.99 & 3.03(AB-System, 1H, J_(AB)=10.1 Hz), 3.15 (m, 1H), 3.73 & 3.84 (AB-System,4H, J_(AB)=14.00), 7.21 (m, 15H); ¹³C NMR (400 MHz, CDCl₃) ∂ 139.55,129.45, 128.42, 128.14, 128.09, 126.84, 125.97, 60.32, 54.23, 52.13,45.99, 33.76; HRMS Calcd for C₂₄H₂₆NO (M+1) 344.477, found 344.2003.

N,N,αS-Tris(phenylmethyl)-2R-oxiranemethanamine

¹H NMR (300 MHz, CDCl₃) ∂ 2.20 (m, 1H), 2.59 (m, 1H), 2.75 (m, 2H), 2.97(m, 1H), 3.14 (m, 1H), 3.85 (AB-System, 4H). 7.25 (m, 15H).HPLC onchiral stationary phase: Pirkle-Whelk-O 1 column (250×4.6 mm I.D.),mobile phase: hexane/isopropanol (99.5:0.5, v/v), flow-rate: 1.5 ml/min,detection with UV detector at 210 nm. Retention time of(8): 9.38 min.,retention time of enantiomer of (4): 13.75 min.

Method 2:

A solution of the crude aldehyde 0.074 mol and chloroiodomethane (7.0ml, 0.096 mol) in tetrahydrofuran (285 ml) was cooled to −78° C., undera nitrogen atmosphere. A 1.6 M solution of n-butyl lithium in hexane (25ml, 0.040 mol) was then added at a rate to maintain the temperature at−75° C. (addition time—15 min.). After the first addition, additionalchloroiodomethane (1.6 ml, 0.022 mol) was added again, followed byn-butyl lithium (23 ml, 0.037 mol), keeping the temperature at −75° C.The mixture was stirred for 15 min. Each of the reagents,chloroiodomethane (0.70 ml, 0.010 mol) and n-butyl lithium (5 ml, 0.008mol) were added 4 more times over 45 min. at −75° C. The cooling bathwas then removed and the solution warmed to 22° C. over 1.5 hr. Themixture was poured into 300 ml of saturated aq. ammonium chloridesolution. The tetrahydrofuran layer was separated. The aqueous phase wasextracted with ethyl acetate (1×300 ml). The combined organic layerswere washed with brine, dried over magnesium sulfate, filtered andconcentrated to give a brown oil (27.4 g). The product could be used inthe next step without purification. The desired diastereomer can bepurified by recrystallization at a subsequent step. The product couldalso be purified by chromatography.

Method 3:

A solution of αS-[Bis(phenylmethyl)amino]benzene-propanaldehyde (178.84g, 0.54 mol) and bromochloromethane (46 mL, 0.71 mol) in tetrahydrofuran(1.8 L) was cooled to −30 to −35° C. (colder temperature such as −70° C.also worked well but warmer temperatures are more readily achieved inlarge scale operations) in a stainless steel reactor under a nitrogenatmosphere. A solution of n-butyl lithium in hexane (1.6 M, 340 mL, 0.54mol) was then added at a rate that maintained the temperature below −25°C. After addition the mixture was stirred at −30 to −35° C. for 10minutes. More additions of reagents were carried out in the followingmanner: (1) additional bromochloromethane (14 mL) was added, followed byn-butyl lithium (102 mL) at <−25° C. After addition the mixture wasstirred at −30 to −35° C. for 10 minutes. This was repeated once. (2)Additional bromochloromethane (7 mL, 0.11 mol) was added, followed byn-butyl lithium (51 mL, 0.082 mol) at <−25° C. After addition themixture was stirred at −30 to −35° C. for 10 minutes. This was repeated5 times. (3) Additional bromochloromethane (7 mL, 0.11 mol) was added,followed by n-butyl lithium (51 mL, 0.082 mol) at <−25° C. Afteraddition the mixture was stirred at −30 to −35° C. for 10 minutes. Thiswas repeated once. The external cooling was stopped and the mixturewarmed to ambient temp. over 4 to 16 hours when TLC (silica gel, 20%ethyl acetate/hexane) indicated that the reaction was completed. Thereaction mixture was cooled to 10° C. and quenched with 1452 g of 16%ammonium chloride solution (prepared by dissolving 232 g of ammoniumchloride in 1220 mL of water), keeping the temperature below 23° C. Themixture was stirred for 10 minutes and the organic and aqueous layerswere separated. The aqueous phase was extracted with ethyl acetate(2×500 mL). The ethyl acetate layer was combined with thetetrahydrofuran layer. The combined solution was dried over magnesiumsulfate (220 g), filtered and concentrated on a rotary evaporator at 65°C. The brown oil residue was dried at 70° C. in vacuo (0.8 bar) for 1 hto give 222.8 g of crude material.

Method 4:

Following the same procedures as described in Method 3 of this Exampleexcept the reaction temperatures were at −20° C. The resultingN,N,αS-tris(phenylmethyl)-2S-oxiranemethanamine was a diastereomericmixture of lesser purity then that of Method 3.

Method 5:

Following the same procedures as described in Method 3 of this Exampleexcept the reaction temperatures were at −70–−78° C. The resultingN,N,αS-tris(phenylmethyl)-2S-oxiranemethanamine was a diastereomericmixture, which was used directly in the subsequent steps withoutpurification.

Method 6:

Following the same procedures as described in Method 3 of this Exampleexcept a continuous addition of bromochloromethane and n-butyl lithiumwas used at −30 to −35° C. After the reaction and work up procedures asdescribed in Method 3 of this Example, the desiredN,N,αS-tris(phenylmethyl)-2S-oxiranemethanamine was isolated incomparable yields and purities.

Method 7:

Following the same procedures as described in Method 2 of this Exampleexcept dibromomethane was used instead of chloroiodomethane. After thereaction and work up procedures as described in Method 2 of thisExample, the desired N,N,αS-tris(phenylmethyl)-2S-oxirane-methanaminewas isolated.

EXAMPLE 4

Preparation ofN-[3(S)-[N,N-bis(phenylmethyl)amino]-2(R)-hydroxy-4-phenylbutyl]-N-isobutylamine

To a solution of crudeN,N-dibenzyl-3(S)-amino-1,2(S)-epoxy-4-phenylbutane (388.5 g, 1.13 mol)in isopropanol (2.7 L) (or ethyl acetate) was added isobutylamine (1.7kgm, 23.1 mol) over 2 min. The temperature increased from 25° C. and to30° C. The solution was heated to 82° C. and stirred at this temperaturefor 1.5 hours. The warm solution was concentrated under reduced pressureat 65° C., The brown oil residue was transferred to a 3-L flask anddried in vacuo (0.8 mm Hg) for 16 h to give 450 g of3S-[N,N-bis(phenylmethyl)amino-4-phenylbutan-2R-ol as a crude oil.

An analytical sample of the desired major diastereomeric product wasobtained by purifying a small sample of crude product by silica gelchromatography (40% ethyl acetate/hexane). Tlc analysis: silica gel, 40%ethyl acetate/hexane; Rf=0.28; HPLC analysis: ultrasphere ODS column,25% triethylamino-/phosphate buffer pH 3-acetonitrile, flow rate 1mL/min, UV detector; retention time 7.49 min.; HRMS Calcd for C₂₈H₂₇N₂O(M+1) 417.616, found 417.2887. An analytical sample of the minordiastereomeric product,3S-[N,N-bis(phenylmethyl)amino]1-(2-methylpropyl)amino-4-phenylbutan-2S-olwas also obtained by purifying a small sample of crude product by silicagel chromatography (40% ethyl acetate/hexane).

EXAMPLE 5

Preparation ofN-[3(S)-[N,N-bis(phenylmethyl)amino]-2(R)-hydroxy-4-phenylbutyl]-N-isobutylamine.oxalicacid salt

To a solution of oxalic acid (8.08 g, 89.72 mmol) in methanol (76 mL)was added a solution of crude3(S)-[N,N-bis(phenylmethyl)amino]-1-(2-methylpropyl)amino-4-phenylbutan-2(R)-ol{39.68 g, which contains about 25.44 g (61.06 mmol) of 3(S),2(R) isomerand about 4.49 g (10.78 mmol) of 3(S),2(S) isomer) in ethyl acetate (90mL) over 15 minutes. The mixture was stirred at room temperature forabout 2 hours. Solid was isolated by filtration, washed with ethylacetate (2×20 mL) and dried in vacuo for about 1 hour to yield 21.86 g(70.7% isomer recovery) of 97% diastereomerically pure salt (based onHPLC peak areas). HPLC analysis: Vydec-peptide/protein C18 column, UVdetector 254 nm, flow rate 2 mL/min., gradient {A=0.05% trifluoroaceticacid in water, B=0.05% trifluoroacetic acid in acetonitrile, 0 min. 75%A/25% B, 30 min. 10% A/90% B, 35 min. 10% A/90% B, 37 min. 75% A/25% B);Retention time 10.68 min. (3(S),2(R) isomer) and 9.73 min. (3(S),2(S)isomer). Mp=174.99° C.; Microanalysis: Calc.: C, 71.05%, H, 7.50%, N,5.53%; Found: C, 71.71%, H, 7.75%, N, 5.39%.

Alternatively, oxalic acid dihydrate (119 g, 0.94 mole) was added to a5000 mL round bottom flask fitted with a mechanical stirrer and adropping funnel. Methanol (1000 ml) was added and the mixture stirreduntil dissolution was complete. A solution of crude3(S)-[N,N-bis(phenylmethyl)amino]-1-(2-methylpropyl)amino-4-phenylbutan-2(R)-olin ethyl acetate (1800 ml, 0.212 g amino alcohol isomers/mL, 0.9160moles) was added over a twenty minute period. The mixture was stirredfor 18 hours and the solid product was isolated by centrifugation in sixportions at 400 G. Each portion was washed with 125 mL of ethyl acetate.The salt was then collected and dried overnight at 1 torr to yield 336.3g of product (71% based upon total amino alcohol). HPLC/MS(electrospray) was consistent with the desired product (m/z 417 [M+H]⁺).

Alternatively, crude3(S)-[N,N-bis(phenylmethyl)amino]-1-(2-methylpropyl)amino-4-phenylbutan-2(R)-ol(5 g) was dissolved in methyl-tert-butylether (MTBE) (10 mL) and oxalicacid (1 g) in methanol (4 mL) was added. The mixture was stirred forabout 2 hours. The resulting solid was filtered, washed with cold MTBEand dried to yield 2.1 g of white solid of about 98.9%diastereomerically pure (based on HPLC peak areas).

EXAMPLE 6 Preparation ofN-[3(S)-[N,N-bis(phenylmethyl)amino]-2(R)-hydroxy-4-phenylbutyl]-N-isobutylamine.aceticacid salt

To a solution of crude 3(S)-[N,N-bis(phenylmethyl)amino]-1-(2-methylpropyl)amino-4-phenylbutan-2(R)-ol inmethyl-tert-butylether (MTBE) (45 mL, 1.1 g amino alcohol isomers/mL)was added acetic acid (6.9 mL) dropwise. The mixture was stirred forabout 1 hour at room temperature. The solvent was removed in vacuo toyield a brown oil about 85% diastereomerically pure product (based onHPLC peak areas). The brown oil was crystallized as follows: 0.2 g ofthe oil was dissolved in the first solvent with heat to obtain a clearsolution, the second solvent was added until the solution became cloudy,the mixture was heated again to clarity, seeded with about 99%diastereomerically pure product, cooled to room temperature and thenstored in a refrigerator overnight. The crystals were filtered, washedwith the second solvent and dried. The diastereomeric purity of thecrystals was calculated from the HPLC peak areas. The results are shownin Table 1.

TABLE 1 First Second Solvent Recovery Diastereomeric Solvent SolventRatio Weight (g) Purity (%) MTBE Heptane  1:10 0.13 98.3 MTBE Hexane 1:10 0.03 99.6 Methanol Water   1:1.5 0.05 99.5 Toluene Heptane  1:100.14 98.7 Toluene Hexane  1:10 0.10 99.7

Alternatively, crude 3(S)-[N,N-bis(phenylmethyl)amino]-1-(2-methylpropyl)amino-4-phenylbutan-2(R)-ol (50.0 g, whichcontains about 30.06 g (76.95 mmol) of 3(S),2(R) isomer and about 5.66 g(13.58 mmol) of 3(S),2(S) isomer) was dissolved inmethyl-tert-butylether (45.0 ML). To this solution was added acetic acid(6.90 mL, 120.6 mmol) over a period of about 10 min. The mixture wasstirred at room temperature for about 1 hour and concentrated underreduced pressure. The oily residue was purified by recrystallizationfrom methyl-tert-butylether (32 mL) and heptane (320 mL). Solid wasisolated by filtration, washed with cold heptane and dried in vacuo forabout 1 hour to afford 21.34 g (58.2% isomer recovery) of 96%diastereomerically pure monoacetic acid salt (based on HPLC peak areas).Mp=105–106° C.; Microanalysis: Calc.: C, 75.53%, H, 8.39%, N, 5.87%;Found: C, 75.05%, H, 8.75%, N, 5.71%.

EXAMPLE 7 Preparation ofN-[3(S)-[N,N-bis(phenylmethyl)amino]-2(R)-hydroxy-4-phenylbutyl]-N-isobutylamine-L-tartaricacid salt

Crude3(S)-[N,N-bis(phenylmethyl)amino]-1-(2-methylpropyl)amino-4-phenylbutan-2(R)-ol(10.48 g, which contains about 6.72 g (16.13 mmol) of 3(S),2(R) isomerand about 1.19 g (2.85 mmol) of 3(S),2(S) isomer) was dissolved intetrahydrofuran (10.0 mL). To this solution was added a solution ofL-tartaric acid (2.85 g, 19 mmol) in methanol (5.0 mL) over a period ofabout 5 min. The mixture was stirred at room temperature for about 10min. and concentrated under reduced pressure. Methyl-tert-butylether(20.0 mL) was added to the oily residue and the mixture was stirred atroom temperature for about 1 hour. Solid was isolated by filtration toafford 7.50 g of crude salt. The crude salt was purified byrecrystallization from ethyl acetate and heptane at room temperature toyield 4.13 g (45.2% isomer recovery) of 95% diastereomerically pureL-tartaric acid salt (based on HPLC peak areas). Microanalysis: Calc.:C, 67.76%, H, 7.41%, N, 4.94%; Found: C, 70.06%, H, 7.47%, N, 5.07%.

EXAMPLE 8 Preparation ofN-[3(S)-[N,N-bis(phenylmethyl)amino]-2(R)-hydroxy-4-phenylbutyl]-N-isobutylamine.dihydrochloricacid salt

Crude 3(S)-[N,N-bis(phenylmethyl)amino]-1-(2-methylpropyl)amino-4-phenylbutan-2(R)-ol (10.0 g, whichcontains about 6.41 g (15.39 mmol) of 3(S),2(R) isomer and about 1.13 g(2.72 mmol) of 3(S),2(S) isomer) was dissolved in tetrahydrofuran (20.0mL). To this solution was added hydrochloric acid (20 mL, 6.0 N) over aperiod of about 5 min. The mixture was stirred at room temperature forabout 1 hour and concentrated under reduced pressure. The residue wasrecrystallized from ethanol at 0° C. to yield 3.20 g (42.7% isomerrecovery) of 98% diastereomerically pure dihydrochloric acid salt (basedon HPLC peak areas). Microanalysis: Calc.: C, 68.64%, H, 7.76%, N,5.72%; Found: C, 68.79%, H, 8.07%, N, 5.55%.

EXAMPLE 9 Preparation ofN-[3(S)-[N,N-bis(phenylmethyl)amino]-2(R)-hydroxy-4-phenylbutyl]-N-isobutylamine.toluenesulfonicacid salt

Crude 3(S)-[N,N-bis(phenylmethyl)amino]-1-(2-methylpropyl)amino-4-phenylbutan-2(R)-ol (5.0 g, whichcontains about 3.18 g (7.63 mmol) of 3(S),2(R) isomer and about 0.56 g(1.35 mmol) of 3(S),2(S) isomer) was dissolved in methyl-tert-butylether(10.0 mL). To this solution was added a solution of toluenesulfonic acid(2.28 g, 12 mmol) in methyl-tert-butylether (2.0 mL) and methanol (2.0mL) over a period of about 5 min. The mixture was stirred at roomtemperature for about 2 hours and concentrated under reduced pressure.The residue was recrystallized from methyl-tert-butylether and heptaneat 0° C., filtered, washed with cold heptane and dried in vacuo to yield1.85 g (40.0% isomer recovery) of 97% diastereomerically puremonotoluenesulfonic acid salt (based on HPLC peak areas).

EXAMPLE 10 Preparation ofN-[3(S)-[N,N-bis(phenylmethyl)amino]-2(R)-hydroxy-4-phenylbutyl]-N-isobutylamine.methanesulfonicacid salt

Crude 3(S)-[N,N-bis(phenylmethyl)amino]-1-(2-methylpropyl)amino-4-phenylbutan-2(R)-ol (10.68 g, whichcontains about 6.85 g (16.44 mmol) of 3(S),2(R) isomer and about 1.21 g(2.90 mmol) of 3(S),2(S) isomer) was dissolved in tetrahydrofuran (10.0mL). To this solution was added methanesulfonic acid (1.25 mL, 19.26mmol). The mixture was stirred at room temperature for about 2 hours andconcentrated under reduced pressure. The oily residue was recrystallizedfrom methanol and water at 0° C., filtered, washed with coldmethanol/water (1:4) and dried in vacuo to yield 2.40 g (28.5% isomerrecovery) of 98% diastereomerically pure monomethanesulfonic acid salt(based on HPLC peak areas).

EXAMPLE 11 Preparation of N-benzyl-L-phenylalaninol

Method 1:

L-Phenylalaninol (89.51 g, 0.592 moles) was dissolved in 375 mL ofmethanol under inert atmosphere, 35.2 g (0.592 moles) of glacial aceticacid and 50 mL of methanol was added followed by a solution of 62.83 g(0.592 moles) of benzaldehyde in 100 mL of methanol. The mixture wascooled to approximately 15° C. and a solution of 134.6 g (2.14 moles) ofsodium cyanoborohydride in 700 mL of methanol was added in approximately40 minutes, keeping the temperature between 15° C. and 25° C. Themixture was stirred at room temperature for 18 hours. The mixture wasconcentrated under reduced pressure and partitioned between 1 L of 2Mammonium hydroxide solution and 2 L of ether. The ether layer was washedwith 1 L of 1M ammonium hydroxide solution, twice with 500 mL water, 500mL of brine and dried over magnesium sulfate for 1 hour. The ether layerwas filtered, concentrated under reduced pressure and the crude solidproduct was recrystallized from 110 mL of ethyl acetate and 1.3 L ofhexane to give 115 g (81% yield) of N-benzyl-L-phenylalaninol as a whitesolid.

Method 2:

L-Phenylalaninol (5 g, 33 mmoles) and 3.59 g (33.83 mmoles) ofbenzaldehyde were dissolved in 55 mL of 3A ethanol under inertatmosphere in a Parr shaker and the mixture was warmed to 60° C. for 2.7hours. The mixture was cooled to approximately 25° C. and 0.99 g of 5%platinum on carbon was added and the mixture was hydrogenated at 60 psiof hydrogen and 40° C. for 10 hours. The catalyst was filtered off, theproduct was concentrated under reduced pressure and the crude solidproduct was recrystallized from 150 mL of heptane to give 3.83 g (48%yield) of N-benzyl-L-phenylalaninol as a white solid.

EXAMPLE 12 Preparation of N-(t-Butoxycarbonyl)-N-benzyl-L-phenylalaninol

N-benzyl-L-phenylalaninol (2.9 g, 12 mmoles) was dissolved in 3 mL oftriethylamine and 27 mL of methanol and 5.25 g (24.1 mmoles) ofdi-tert-butyl dicarbonate was added. The mixture was warmed to 60° C.for 35 minutes and concentrated under reduced pressure. The residue wasdissolved in 150 mL of ethyl acetate and washed twice with 10 mL of cold(0–5° C.), dilute hydrochloric acid (pH 2.5 to 3), 15 mL of water, 10 mLof brine, dried over magnesium sulfate, filtered and concentrated underreduced pressure. The crude product oil was purified by silica gelchromatography (ethyl acetate: hexane, 12:3 as eluting solvent) to give3.98 g (97% yield) of colorless oil.

EXAMPLE 13 Preparation of N-(t-Butoxycarbonyl)-N-benzyl-L-phenylalaninal

Method 1:

To a solution of 0.32 g (0.94 mmoles) ofN-(t-butoxycarbonyl)-N-benzyl-L-phenylalaninol in 2.8 mL of toluene wasadded 2.4 mg (0.015 mmoles) of 2,2,6,6-tetramethyl-1-piperidinyloxy,free radical (TEMPO), 0.1 g (0.97 mmoles) of sodium bromide, 2.8 mL ofethyl acetate and 0.34 mL of water. The mixture was cooled to 0° C. andan aqueous solution of 4.2 mL of 5% household bleach containing 0.23 g(3.0 mL, 2.738 mmoles) of sodium bicarbonate was added slowly over 30minutes. The mixture was stirred at 0° C. for 10 minutes. Three moreadditions (0.4 mL each) of bleach was added followed by stirring for 10minutes after each addition to consume all the stating material. The twophase mixture was allowed to separate. The aqueous layer was extractedtwice with 8 mL of toluene. The combined organic layer was washed with1.25 mL of a solution containing 0.075 g of potassium iodide, sodiumbisulfate(0.125 g) and water(1.1 mL), 1.25 mL of 10% aqueous sodiumthiosulfate solution, 1.25 mL of pH 7 phosphate buffer and 1.5 mL ofbrine solution. The organic solution was dried over magnesium sulfate,filtered and concentrated under reduced pressure to give 0.32 g (100%yield) of N-(t-Butoxycarbonyl)-N-benzyl-L-phenylalaninal.

Method 2:

To a solution of 2.38 g(6.98 mmoles) ofN-(t-butoxycarbonyl)-N-benzyl-L-phenylalaninol in 3.8 mL (27.2 mmoles)of triethylamine at 10° C. was added a solution of 4.33 g (27.2 mmoles)of sulfur trioxide pyridine complex in 17 mL of dimethyl sulfoxide. Themixture was warmed to room temperature and stirred for one hour. Water(16 mL) was added and the mixture was extracted with 20 mL of ethylacetate. The organic layer was washed with 20 mL of 5% citric acid, 20mL of water, 20 mL of brine, dried over magnesium sulfate and filtered.The filtrate was concentrated under reduced pressure to give 2.37 g(100% yield) of N-(t-Butoxycarbonyl)-N-benzyl-L-phenylalaninal.

EXAMPLE 14

Preparation of3(S)-[N-(t-butoxycarbonyl)-N-benzylamino]-1,2-(S)-epoxy-4-phenylbutane

Method 1:

A solution of 2.5 g (7.37 mmoles) ofN-(t-butoxycarbonyl)-N-benzyl-L-phenylalaninal and 0.72 mL ofchloroiodomethane in 35 mL of THF was cooled to −78° C. A 4.64 mL of asolution of n-butyllithium (1.6 M in hexane, 7.42 mmoles) was addedslowly, keeping the temperature below −70° C. The mixture was stirredfor 10 minutes between −70 to −75° C. Two additional portions of 0.22 mLof chloroiodomethane and 1.4 mL of n-butyllithium was added sequentiallyand the mixture was stirred for 10 minutes between −70 to −75° C. aftereach addition. Four additional portions of 0.11 mL of chloroiodomethaneand 0.7 mL of n-butyllithium was added sequentially and the mixture wasstirred for 10 minutes between −70 to −75° C. after each addition. Themixture was warmed to room temperature for 3.5 hours. The product wasquenched at below 5° C. with 24 mL of ice-cold water. The biphasiclayers were separated and the aqueous layer was extracted twice with 30mL of ethyl acetate. The combined organic layers was washed three timeswith 10 mL water, then with 10 mL brine, dried over sodium sulfate,filtered and concentrated under reduced pressure to give 2.8 g of ayellow crude oil. This crude oil (>100% yield) is a mixture of thediastereomeric epoxidesN,αS-bis(phenylmethyl)-N-(t-butoxycarbonyl)-2S-oxiranemethanamine andN,αS-bis(phenylmethyl)-N-(t-butoxycarbonyl)-2R-oxiranemethanamine. Thecrude mixture is used directly in the next step without purification.

Method 2:

To a suspension of 2.92 g (13.28 mmoles) of trimethylsulfoxonium iodidein 45 mL of acetonitrile was added 1.49 g (13.28 mmoles) of potassiumt-butoxide. A solution of 3.0 g (8.85 mmoles) ofN-(t-butoxycarbonyl)-N-benzyl-L-phenylalaninal in 18 mL of acetonitrilewas added and the mixture was stirred at room temperature for one hour.The mixture was diluted with 150 mL of water and extracted twice with200 mL of ethyl acetate. The organic layers were combined and washedwith 100 mL water, 50 mL brine, dried over sodium sulfate, filtered andconcentrated under reduced pressure to give 3.0 g of a yellow crude oil.The crude product was purified by silica gel chromatography (ethylacetate/hexane: 1:8 as eluting solvent) to give 1.02 g (32.7% yield) ofa mixture of the two diastereomersN,αS-bis(phenylmethyl)-N-(t-butoxycarbonyl)-2S-oxiranemethanamine andN,αS-bis(phenylmethyl)-N-(t-butoxycarbonyl)-2R-oxiranemethanamine.

Method 3:

To a suspension of 0.90 g (4.42 mmoles) of trimethylsulfonium iodide in18 mL of acetonitrile was added 0.495 g (4.42 mmoles) of potassiumt-butoxide. A solution of 1.0 g (2.95 mmoles) ofN-(t-butoxycarbonyl)-N-benzyl-L-phenylalaninal in 7 mL of acetonitrilewas added and the mixture was stirred at room temperature for one hour.The mixture was diluted with 80 mL of water and extracted twice with 80mL of ethyl acetate. The organic layers were combined and washed with100 mL water, 30 mL brine, dried over sodium sulfate, filtered andconcentrated under reduced pressure to give 1.04 g of a yellow crudeoil. The crude product was a mixture of the two diastereomersN,αS-bis(phenylmethyl)-N-(t-butoxycarbonyl)-2S-oxiranemethanamine andN,αS-bis(phenylmethyl)-N-(t-butoxycarbonyl)-2R-oxiranemethanamine.

EXAMPLE 15

Preparation of 3S-[N-(t-Butoxycarbonyl)-N-(phenylmethyl)amino]-1-(2-methylpropyl)amino-4-phenylbutan-2R-ol

To a solution of 500 mg (1.42 mmoles) of the crude epoxide (a mixture ofthe two diastereomersN,αS-bis(phenylmethyl)-N-(t-butoxycarbonyl)-2S-oxiranemethanamine andN,αS-bis(phenylmethyl)-N-(t-butoxycarbonyl)-2R-oxiranemethanamine) in0.98 mL of isopropanol was added 0.71 mL (7.14 mmoles) of isobutylamine.The mixture was warmed to reflux at 85° C. to 90° C. for 1.5 hours. Themixture was concentrated under reduced pressure and the product oil waspurified by silica gel chromatography (chloroform:methanol, 100:6 aseluting solvents) to give 330 mg of3S-[N-(t-butoxycarbonyl)-N-(phenylmethyl)amino]-1(2-methylpropyl)amino-4-phenylbutan-2R-olas a colorless oil (54.5% yield).3S-[N-(t-Butoxycarbonyl)-N-(phenylmethyl)amino]-1-(2-methylpropyl)amino-4-phenylbutan-2S-olwas also isolated. When purifiedN,αS-bis(phenylmethyl)-N-(t-butoxycarbonyl)-2S-oxiranemethanamine wasused as starting material,3S-[N-(t-butoxycarbonyl)-N-(phenylmethyl)amino]-1(2-methylpropyl)amino-4-phenylbutan-2R-olwas isolated after purification by chromatography in an 86% yield.

EXAMPLE 16

Preparation of 3S-(N-t-Butoxycarbonyl)amino-4-phenylbutan-1,2R-diol

To a solution of 1 g (3.39 mmoles) of2S-(N-t-butoxycarbonyl)amino-1S-hydroxy-3-phenylbutanoic acidcommercially available from Nippon Kayaku, Japan) in 50 mL of THF at 0°C. was added 50 mL of borane-THF complex (liquid, 1.0 M in THF), keepingthe temperatures below 5° C. The reaction mixture was warmed to roomtemperature and stirred for 16 hours. The mixture was cooled to 0° C.and 20 mL of water was added slowly to destroy the excess BH₃ and toquench the product mixture, keeping the temperature below 12° C. Thequenched mixture was stirred for 20 minutes and concentrated underreduced pressure. The product mixture was extracted three times with 60mL of ethyl acetate. The organic layers were combined and washed with 20mL of water, 25 mL of saturated sodium chloride solution andconcentrated under reduced pressure to give 1.1 g of crude oil. Thecrude product was purified by silica gel chromatography(chloroform/methanol, 10:6 as eluting solvents) to give 900 mg (94.4%yield) of 3S-(N-t-butoxycarbonyl)amino-4-phenylbutan-1,2R-diol as awhite solid.

EXAMPLE 17

Preparation of 3S-(N-t-Butoxycarbonyl)amino-2R-hydroxy-4-phenylbutylToluenesulfonate

To a solution of 744.8 mg (2.65 mmoles) of3S-(N-t-butoxycarbonyl)amino-4-phenylbutan-1,2R-diol in 13 mL ofpyridine at 0° C. was added 914 mg of toluenesulfonyl chloride in oneportion. The mixture was stirred at 0° C. to 5° C. for 5 hours. Amixture of 6.5 mL of ethyl acetate and 15 mL of 5% aqueous sodiumbicarbonate solution was added to the reaction mixture and stirred for 5minutes. The product mixture was extracted three times with 50 mL ofethyl acetate. The organic layers were combined and washed with 15 mL ofwater, 10 mL of saturated sodium chloride solution and concentratedunder reduced pressure to give about 1.1 g of a yellow chunky solid. Thecrude product was purified by silica gel chromatography (ethylacetate/hexane 1:3 as eluting solvents) to give 850 mg (74% yield) of3S-(N-t-butoxycarbonyl)amino-2R-hydroxy-4-phenylbut-1-yltoluenesulfonate as a white solid.

EXAMPLE 18

Preparation of3S-[N-(t-Butoxycarbonyl)amino]-1-(2-methylpropyl)amino-4-phenylbutan-2R-ol

To a solution of 90 mg (0.207 mmoles) of3S-(N-t-butoxycarbonyl)amino-2R-hydroxy-4-phenylbut-1-yltoluenesulfonate in 0.143 mL of isopropanol and 0.5 mL of toluene wasadded 0.103 mL (1.034 mmoles) of isobutylamine. The mixture was warmedto 80 to 85° C. and stirred for 1.5 hours. The product mixture wasconcentrated under reduced pressure at 40 to 50° C. and purified bysilica gel chromatography (chloroform/methanol, 10:1 as elutingsolvents) to give 54.9 mg (76.8% yield) of3S-[N-(t-butoxycarbonyl)amino]-1-(2-methylpropyl)amino-4-phenylbutan-2R-olas a white solid.

EXAMPLE 19

Preparation ofN-[3(S)-benzyloxycarbonylamino-2(R)-hydroxy-4-phenylbutyl]-N-isobutylamine

Part A:

To a solution of 75.0 g (0.226 mol) ofN-benzyloxycarbonyl-L-phenylalanine chloromethyl ketone in a mixture of807 mL of methanol and 807 mL of tetrahydrofuran at −2° C., was added13.17 g (0.348 mol, 1.54 equiv.) of solid sodium borohydride over onehundred minutes. The solvents were removed under reduced pressure at 40°C. and the residue dissolved in ethyl acetate (approx. 1 L). Thesolution was washed sequentially with 1M potassium hydrogen sulfate,saturated sodium bicarbonate and then saturated sodium chloridesolutions. After drying over anhydrous magnesium sulfate and filtering,the solution was removed under reduced pressure. To the resulting oilwas added hexane (approx. 1 L) and the mixture warmed to 60° C. withswirling. After cooling to room temperature, the solids were collectedand washed with 2 L of hexane. The resulting solid was recrystallizedfrom hot ethyl acetate and hexane to afford 32.3 g (43% yield) ofN-benzyloxycarbonyl-3(S)-amino-1-chloro-4-phenyl-2(S)-butanol, mp150–151° C. and M+Li⁺=340.

Part B:

To a solution of 6.52 g (0.116 mol, 1.2 equiv.) of potassium hydroxidein 968 mL of absolute ethanol at room temperature, was added 32.3 g(0.097 mol) of N-CBZ-3(S)-amino-1-chloro-4-phenyl-2(S)-butanol. Afterstirring for fifteen minutes, the solvent was removed under reducedpressure and the solids dissolved in methylene chloride. After washingwith water, drying over magnesium sulfate, filtering and stripping, oneobtains 27.9 g of a white solid. Recrystallization from hot ethylacetate and hexane afforded 22.3 g (77% yield) ofN-benzyloxycarbonyl-3(S)-amino-1,2(S)-epoxy-4-phenylbutane, mp 102–103°C. and ME⁺ 298.

Part C:

A solution of N-benzyloxycarbonyl3(S)-amino-1,2-(S)-epoxy-4-phenylbutane (1.00 g, 3.36 mmol) andisobutylamine (4.90 g, 67.2 mmol, 20 equiv.) in 10 mL of isopropylalcohol was heated to reflux for 1.5 hours. The solution was cooled toroom temperature, concentrated in vacuo and then poured into 100 mL ofstirring hexane whereupon the product crystallized from solution. Theproduct was isolated by filtration and air dried to give 1.18 g, 95% ofN-[[3(S)-phenylmethylcarbamoyl)amino-2(R)-hydroxy-4-phenylbutyl]N-[(2-methylpropyl)]amine,C₂₂H₃₀N₂O₃, mp 108.0–109.5° C., MH⁺ m/z=371.

EXAMPLE 20

Preparation of phenylmethyl [2R-hydroxy-3-[(3-methylbutyl)(phenylsulfonyl)amino]-1S-(phenylmethyl)propyl]carbamate

From the reaction ofN[³(S)-benzyloxycarbonylamino-2(R)-hydroxy-4-phenylbutyl]N-isoamylamine(1.47 gm, 3.8 mmol), triethylamine (528 uL, 3.8 mmol) andbenzenesulfonyl chloride (483 uL, 3.8 mmol) one obtains phenylmethyl[2R-hydroxy-3-[(3-methylbutyl)(phenylsulfonyl)amino]-1S-(phenylmethyl)propyl]-carbamate. Columnchromatography on silica gel eluting with chloroform containing 1%ethanol afforded the pure product. Anal. Calcd for C₂₉H₃₆N₂O₅S: C,66.39; H, 6.92; N, 5.34. Found: C, 66.37; H, 6.93; N, 5.26.

EXAMPLE 21

Preparation of2R-hydroxy-3-[[(4-aminophenyl)sulfonyl](2-methylpropyl)amino]-1S-(phenylmethyl)propylamine

Part A: Preparation of Carbamic acid,2R-hydroxy-3-[[(4-nitrophenyl)sulfonyl](2-methylpropyl)amino]-1S-(phenylmethyl)propyl-,phenylmethyl ester

To a solution of 4.0 g (10.8 mmol) of N-[3S-benzyloxycarbonylamino-2R-hydroxy-4-phenyl]-N-isobutylamine in 50 mL of anhydrousmethylene chloride, was added 4.5 mL (3.27 g, 32.4 mmol) oftriethylamine. The solution was cooled to 0° C. and 2.63 g (11.9 mmol)of 4-nitrobenzene sulfonyl chloride was added, stirred for 30 minutes at0° C., then for 1 hour at room temperature. Ethyl acetate was added,washed with 5% citric acid, saturated sodium bicarbonate, brine, driedand concentrated to yield 5.9 g of crude material. This wasrecrystallized from ethyl acetate/hexane to afford 4.7 g of purecarbamic acid,[2R-hydroxy-3-[[(4-nitrophenyl)sulfonyl](2-methylpropyl)amino]-1S-(phenylmethyl)propyl-,phenylmethyl ester, m/e=556(M+H).

Part B: Preparation of2R-hydroxy-3-[[(4-aminophenyl)sulfonyl](2-methylpropyl)amino]-1S-(phenylmethyl)propylamine

A solution of 3.0 g (5.4 mmol) of carbamic acid,2R-hydroxy-3-[[(4-nitrophenyl)sulfonyl](2-methylpropyl)amino]-1S-(phenylmethyl)propyl-, phenylmethyl ester in 20 mL of ethylacetate was hydrogenated over 1.5 g of 10% palladium-on-carbon catalystunder 35 psig of hydrogen for 3.5 hours. The catalyst was removed byfiltration and the solution concentrated to afford 2.05 g of the desired2R-hydroxy-3-[[(4-aminophenyl)sulfonyl](2-methylpropyl)amino]-1S-(phenylmethyl)propylamine,m/e=392(M+H).

EXAMPLE 22

Preparation of2R-hydroxy-3-[[(3-aminophenyl)sulfonyl](2-methylpropyl)amino]-1S-(phenylmethyl)propylamine

Part A: Preparation of Carbamic acid,[2R-hydroxy-3-[(3-nitrophenylsulfonyl)(2-methylpropyl)amino]-1S-(phenylmethyl)propyl-,phenylmethyl ester

To a solution of 1.1 g (3.0 mmol) of N-(3S-benzyloxycarbonylamino-2R-hydroxy-4-phenyl]-N-isobutylamine in 15 mL of anhydrousmethylene chloride, was added 1.3 mL (0.94 g, 9.3 mmol) oftriethylamine. The solution was cooled to 0° C. and 0.67 g (3.0 mmol) of3-nitrobenzene sulfonyl chloride was added, stirred for 30 minutes at 0°C., then for 1 hour at room temperature. Ethyl acetate was added, washedwith 5% citric acid, saturated sodium bicarbonate, brine, dried andconcentrated to yield 1.74 g of crude material. This was recrystallizedfrom ethyl acetate/hexane to afford 1.40 g of pure carbamic acid,[2R-hydroxy-3-[(3-nitrophenylsulfonyl)(2-methylpropyl)amino]-1S-(phenylmethyl)propyl-, phenylmethyl ester, m/e=562(M+Li).

Part B: Preparation of[2R-hydroxy-3-[[(3-aminophenyl)sulfonyl](2-methylpropyl)amino]-1S-(phenylmethyl)propylamine

A solution of 1.33 g (2.5 mmol) of carbamic acid,[2R-hydroxy-3-[(3-nitrophenylsulfonyl)(2-methylpropyl)amino]-1S-(phenylmethyl)propyl-,phenylmethyl ester in 40 mL of 1:1 methanol/tetrahydrofuran washydrogenated over 0.70 g of 10% palladium-on-carbon catalyst under 40psig of hydrogen for 1.5 hours. The catalyst was removed by filtrationand the solution concentrated to afford 0.87 g of the desired[2R-hydroxy-3-[[(3-aminophenyl)sulfonyl](2-methylpropyl)amino]-1S-(phenylmethyl)propylamine.

EXAMPLE 23

Preparation of2R-hydroxy-3-[[(2,3-dihydrobenzofuran-5-yl)sulfonyl](2-methylpropyl)amino]-1S-(phenylmethyl)propylamine

Part A: Preparation of 5-(2,3-dihydrobenzofuranyl) sulfonyl chloride

To a solution of 3.35 g of anhydrous N,N-dimethylformamide at 0° C.under nitrogen was added 6.18 g of sulfuryl chloride, whereupon a solidformed. After stirring for 15 minutes, 4.69 g of 2,3-dihydrobenzofuranwas added, and the mixture heated at 100° C. for 2 hours. The reactionwas cooled, poured into ice water, extracted with methylene chloride,dried over magnesium sulfate, filtered and concentrated the crudematerial. This was recrystallized from ethyl acetate to afford 2.45 g of5-(2,3-dihydrobenzofuranyl)sulfonyl chloride.

Part B: Preparation of Carbamic acid,2R-hydroxy-3-[[(2,3-dihydrobenzofuran-5-yl)sulfonyl](2-methylpropyl)amino]-1S-(phenylmethyl)propyl-,phenylmethyl ester

To a solution of 1.11 g (3.0 mmol) of N-[3S-benzyloxycarbonylamino-2R-hydroxy-4-phenyl]-N-isobutylamine in 20 mL of anhydrousmethylene chloride, was added 1.3 mL (0.94 g, 9.3 mmol) oftriethylamine. The solution was cooled to 0° C. and 0.66 g of5-(2,3-dihydrobenzofuranyl) sulfonyl chloride was added, stirred for 15minutes at 0° C., then for 2 hour at room temperature. Ethyl acetate wasadded, washed with 5% citric acid, saturated sodium bicarbonate, brine,dried and concentrated to yield 1.62 g of crude material. This wasrecrystallized from diethyl ether to afford 1.17 g of pure carbamicacid,[2R-hydroxy-3-[[(2,3-dihydrobenzofuran-5-yl)sulfonyl](2-methylpropyl)amino]-1S-(phenylmethyl)propyl-,phenylmethyl ester

Part C: Preparation of [2R-hydroxy-3-[[(2,3-dihydrobenzofuran-5-yl)sulfonyl)(2-methylpropyl)amino]-1S-(phenylmethyl)propylamine

A solution of 2.86 g of carbamic acid,[2R-hydroxy-3-[[(2,3-dihydrobenzofuran-5-yl)sulfonyl](2-methylpropyl)amino]-1S-(phenylmethyl)propyl-,phenylmethyl ester in 30 mL of tetrahydrofuran was hydrogenated 0.99 gof 10% palladium-on-carbon under 50 psig of hydrogen for 16 hours. Thecatalyst was removed by filtration and the filtrate concentrated toafford 1.99 g of the desired[2R-hydroxy-3-[[(2,3-dihydrobenzofuran-5-yl)sulfonyl](2-methylpropyl)amino]-1S-(phenylmethyl)propylamine.

EXAMPLE 24

Preparation ofN-[(1,1-dimethylethoxyl)carbonyl]-N-[2-methylpropyl]-3S-[N¹-(phenylmethoxycarbonyl)amino]-2R-hydroxy-4-phenylbutylamine

To a solution of 7.51 g (20.3 mmol) ofN-[3S-[(phenylmethoxycarbonyl)amino]-2R-hydroxy-4-phenylbutyl)-2-methylpropylaminein 67 mL of anhydrous tetrahydrofuran was added 2.25 g (22.3 mmol) oftriethylamine. After cooling to 0° C., 4.4 g (20.3 mmol) ofdi-tert-butyldicarbonate was added and stirring continued at roomtemperature for 21 hours. The volatiles were removed in vacuo, ethylacetate added, then washed with 5% citric acid, saturated sodiumbicarbonate, brine, dried over magnesium sulfate, filtered andconcentrated to afford 9.6 g of crude product. Chromatography on silicagel sing 30% ethyl acetate/hexane afforded 8.2 g of pureN-[[3S-(phenylmethylcarbamoyl)amino]-2R-hydroxy-4-phenyl]-1-[(2-methylpropyl)amino-2-(1,1-dimethylethoxyl)carbonyl]butane,mass spectum m/e=477 (M+Li).

EXAMPLE 25

Preparation of 2-methyl-3-[(2-phenylethyl)sulfonyl]propionic acidN-hydroxybenzotriazole ester

Part A: A solution of methyl methacrylate (7.25 g, 72.5 mmol) andphenethyl mercaptan (10.0 g, 72.5 mmol) in 100 mL of methanol was cooledin an ice bath and treated with sodium methoxide (100 mg, 1.85 mmol).The solution was stirred under nitrogen for 3 h and then concentrated invacuo to give an oil that was taken up in ether and washed with 1 Naqueous potassium hydrogen sulfate, saturated aqueous sodium chloride,dried over anhydrous magnesium sulfate, filtered and concentrated togive 16.83 g, 97.5% of methyl 2-(R,S)-methyl-4-thia-6-phenyl hexanoateas an oil. TLC on SiO₂ eluting with 20:1 hexane:ethyl acetate (v:v)R_(f)=0.41. Alternatively, one can use methyl 3-bromo-2-methylpropionate in place of methyl methacrylate.

Part B: A solution of methyl 2-(R,S)-methyl-4-thia-6-phenyl hexanoate(4.00 g, 16.8 mmol) in 100 mL of dichloromethane was stirred at roomtemperature and treated portion wise with meta-chloroperoxybenzoic acid(7.38 g, 39.2 mmol) over approximately 40 m. The solution was stirred atroom temperature for 16 h and then filtered and the filterate washedwith saturated aqueous sodium bicarbonate, 1N sodium hydroxide,saturated aqueous sodium chloride, dried over anhydrous magnesiumsulfate, filtered, and concentrated to give 4.50 g, 99% of desiredsulfone. The unpurified sulfone was dissolved in 100 mL oftetrahydrofuran and treated with a solution of lithium hydroxide (1.04g, 24.5 mmol) in 40 mL of water. The solution was stirred at roomtemperature for 2 m and then concentrated in vacuo. The residue was thenacidified with 1N aqueous potassium hydrogen sulfate to pH=1 and thenextracted three times with ethyl acetate. The combined ethyl acetatesolution was washed with saturated aqueous sodium chloride, dried overanhydrous magnesium sulfate, filtered and concentrated to give a whitesolid. The solid was taken up in boiling ethyl acetate/hexane andallowed to stand undisturbed whereupon white needles formed that wereisolated by filtration and air dried to give 3.38 g, 79% of2-(R,S)-methyl-3 (β-phenethylsulfonyl)-propionic acid, mp 91–93° C.

Part C: A solution of 2-(R,S)-methyl-3(β-phenethylsulfonyl)-propionicacid (166.1 mg, 0.65 mmol), N-hydroxybenzotriazole (HOBT) (146.9 mg,0.97 mmol), and 1-(3-dimethylaminopropyl)-3-ethylcarbodiimidehydrochloride (EDC) (145.8 mg, 0.75 mmol) in 4 mL of anhydrousdimethylformamide (DMF) cooled to 0° C. and stirred under nitrogen for0.5 h. This solution is then treated with a desired protected amino orsulfonamide isostere intermediate and stirred at room temperature for 16h. The solution is poured into 30 mL of 60% saturated aqueous sodiumbicarbonate solution. The aqueous solution is then decanted from theorganic residue. The organic residue is taken up in dichloromethane andwashed with 10% aqueous citric acid, brine, dried over anhydrousmagnesium sulfate, filtered and concentrated. Flash chromatography ofthe mixture on silica gel eluting with 1:1 hexane:ethyl acetate can beutilized and will afford the separated diastereomers.

EXAMPLE 26

Preparation of 2-methyl-3-(methylsulfonyl)propionic acidN-hydroxybenzotriazole ester

Part A: A solution of methyl 2-(bromomethyl)-acrylate (26.4 g, 0.148mol) in 100 mL of methanol was treated with sodium methanesulfinate(15.1 g, 0.148 mol) portion wise over 10 m at room temperature. Thesolution was then stirred at room temperature for a period of 1.25 h andthe solution concentrated in vacuo. The residue was then taken up inwater and extracted four times with ethyl acetate. The combined ethylacetate solution was washed with saturated sodium chloride, dried overanhydrous magnesium sulfate, filtered and concentrated to give a whitesolid, 20.7 g which was taken up in boiling acetone/methyl tert-butylether and allowed to stand whereupon crystals of pure methyl2-(methylsulfonylmethyl) acrylate 18.0 g, 68% formed, mp 65–68° C.

Part B: A solution of methyl 2-(methylsulfonylmethyl) acrylate (970 mg,5.44 mmol) in 15 mL of tetrahydrofuran was treated with a solution oflithium hydroxide (270 mg, 6.4 mmol) in 7 mL of water. The solution wasstirred at room temperature for 5 m and then acidified to pH=1 with 1 Naqueous potassium hydrogen sulfate and the solution extracted threetimes with ethyl acetate. The combined ethyl acetate solution was driedover anhydrous magnesium sulfate, filtered, and concentrated to give 793mg, 89% of 2-(methylsulfonylmethyl) acrylic acid, mp 147–149° C.

Part C: A solution of 2-(methylsulfonylmethyl) acrylic acid (700 mg,4.26 mmol) in 20 mL of methanol was charged into a Fisher-Porter bottlealong with 10% palladium on carbon catalyst under a nitrogen atmosphere.The reaction vessel was sealed and flushed five times with nitrogen andthen five times with hydrogen. The pressure was maintained at 50 psigfor 16 h and then the hydrogen was replaced with nitrogen and thesolution filtered through a pad of celite to remove the catalyst and thefilterate concentrated in vacuo to give 682 mg 96% of2-(R,S)-methyl-3-methylsulfonyl propionic acid.

Part D: A solution of 2-(R,S)-methyl-3(methylsulfonyl) propionic acid(263.5 mg, 1.585 mmol), N-hydroxybenzotriazole (HOBT) (322.2 mg, 2.13mmol), and 1-(3-dimethylaminopropyl)-3-ethylcarbodiimide hydrochloride(EDC) (339.1 mg, 1.74 mmol) in 4 mL of anhydrous dimethylformamide (DMF)is cooled to 0° C. and stirred under nitrogen for 0.5 h. This solutionis then treated with a desired protected amino or sulfonamide isostereintermediate and stirred at room temperature for 16 h. The solution ispoured into 60 mL of 60% saturated aqueous sodium bicarbonate solution.The aqueous solution is then decanted from the organic residue. Theorganic residue is taken up in dichloromethane and washed with 10%aqueous citric acid, brine, dried over anhydrous magnesium sulfate,filtered and concentrated to give the desired product.

EXAMPLE 26A

Preparation of N-[2R-hydroxy-3-[[(1,1-dimethylethoxy)carbonyl](2-methylpropyl)amino]-1s-(phenylmethyl)propyl]-2-R,S-methyl-3-(methylsulfonyl)propanamide

Part A:N-[(1,1-dimethylethoxyl)carbonyl]-N-[2-methylpropyl]-3S-[N¹-(phenylmethoxycarbonyl)amino]-2R-hydroxy-4-phenylbutylamine

from Example 24 dissolved in ethanol was hydrogenated at 45 psi ofhydrogen in the presence of 5% Pd(C) catalyst to yieldN-[(1,1-dimethylethoxyl)carbonyl]-N-[2-methylpropyl]-3S-[N¹-amino]-2R-hydroxy-4-phenylbutylamine.Following standard workup by filtration of the 5% Pd(C) catalyst andevaporation of the filtrate solvent under reduced pressure using arotary evaporator, the amine was obtained

Part B:

The amine from Part A is reacted in DMF with2-methyl-3-(methylsulfonyl)propionic acid N-hydroxybenzotriazole esterfrom Example 26 at or about room temperature. The solution is washedwith sodium bicarbonate solution and extracted with ethyl acetate. Theethyl acetate extract is washed with citric acid solution, brine and isdried over sodium sulfate. The drying agent is filtered and the organicsolvent is removed to provide product

EXAMPLE 26B

Preparation ofN-[2R-hydroxy-3-[(2-methylpropyl)[1,3-benzodioxol-5-yl)sulfonyl]amino]-1S-(phenylmethyl)propyl]-3S-[2-R,S-methyl-3-(methylsulfonyl)]propanamide

N-[2R-hydroxy-3-[[(1,1-dimethylethoxy)carbonyl](2-methylpropyl)amino]-1S-(phenylmethyl)propyl]-3S-[2-R,S-methyl-3-(methylsulfonyl)]propanamide(Example 26A) is dissolved in dioxane/HCl and it is stirred for about 2hours at room temperature. The solvent is removed and the residue isdried in vacuo to produce the amine

The residue is stirred in ethyl acetate, 1,3-benzodioxol-5-yl sulfonylchloride is added followed by triethylamine and the mixture is stirredat about room temperature. The reaction mixture is diluted with ethylacetate, is washed with saturated sodium bicarbonate (saturated) andbrine, dried (MgSO₄) and concentrated to provide product.

The residue is chromatographed if further purification and/or separationof the isomer (e.g., see below) is desired.

EXAMPLE 27 Preparation of Sulfone Inhibitors FromL-(+)-S-acetyl-β-mercaptoisobutyric Acid

Part A: A round-bottomed flask is charged with the desired protectedamino or sulfonamide isostere intermediate (2.575 mmol) and coupled toL-(+)-S-acetyl-b-mercapto butyric acid in the presence of1-(3-dimethylaminopropyl)-3-ethylcarbodiimide hydrochloride (EDC) (339.1mg, 1.74 mmol), in 10 mL of CH₂Cl₂ and is allowed to stir at roomtemperature for 16 h. The solution is concentrated in vacuo and theresidue taken up in ethyl acetate, washed with 1N KHSO₄ sat. aq. NaHCO₃,brine, dried over anhydrous MgSO₄, filtered and concentrated to give anoil which can be purified by radial chromatography on SiO₂ eluting withethyl acetate to give the pure product.

Part B: A solution of the product of Part A (0.85 mmol) in 10 mL ofmethanol is treated with anhydrous ammonia for ca. 1 m at 0° C. Thesolution is stirred at that temperature for 16 h and then concentratedin vacuo to give the desired product that can be used directly in thenext step without further purification.

Part C: A solution of the product of Part B (0.841 mmol) in 10 mL of drytoluene under nitrogen is treated in rapid succession with1,8-diazabicyclo[5.4.0]undec-7-ene, (DBU), (128.1 mg. 0.841 mmol) andiodomethane (119.0 mg, 0.841 mmol). After 0.5 h at room temperature thereaction is diluted with ethyl acetate washed with 1N KHSO₄, sat. aq.NaHCO₃, brine. After the solution is dried over anhydrous MgSO₄,filtered and concentrated in vacuo the desired product is obtained andcan be used directly in the next step.

Part D: A solution of the product of Part C (0.73 mmol) and sodiumperborate (500 mg, 3.25 mmol) in 30 mL of glacial acetic acid is warmedto 55° C. for 16 h. The solution is conentrated in vacuo and then theresidue is taken up in ethyl acetate, washed with water, sat. aq.NaHCO₃, brine, dried over anhydrous MgSO₄, filtered and concentrated togive the desired product.

General Procedure for Coupling Sulfonyl Compounds to Sulfonamides

A mixture of the sulfonyl alkanoyl compound (approximately 1 mmol),N-hydroxybenzotriazole (1.5 mmol), and1-(3-dimethylaminopropyl)-3-ethylcarbodiimide hydrochloride (EDC) (1.2mmol) is dissolved in a suitable solvent such as DMF and allowed toreact for about 30 min. at 0° C. A desired protected amino orsulfonamide isostere intermediate (1.05 mmol) is dissolved in DMF, addedto the above mixture and stirred at room temperature for a period oftime sufficient for the reaction to take place. The solution is thenpoured into saturated aqueous NaHCO₃ and extracted with, for example,ethyl acetate. The extracts are washed, dried, filtered andconcentrated. The resulting material is then crystallized from asuitable solvent or solvent mixture such as hexanes and ethyl acetate toproduce the product.

EXAMPLE 28

Preparation of 2(S)-methyl-3-(methylsulfonyl)propionic Acid

Part A: To a solution of 10 g of D-(−)-S-benzoyl-b-mercaptioisobutyricacid t-butyl ester in 20 mL of methanol was bubbled in gaseous ammoniaat 0° C. The reaction was allowed to then warm to room temperature,stirred overnight and concentrated under reduced pressure. The resultingmixture of a solid (benzamide) and liquid was filtered to provide 5.21 gof a pale oil which then solidified. This was identified as2(S)-methyl-3-mercaptopropionic acid t-butyl ester.

Part B: To a solution of 5.21 g of 2(S)-methyl-3-mercaptopropionic acidt-butyl ester in 75 mL of toluene at 0° C. was added 4.50 g of1,8-diazabicyclo[5.40]undec-7-ene and 1.94 mL of methyl iodide. Afterstirring at room temperature for 2.5 hours, the volatiles were removed,ethyl acetate added, washed with dilute hydrochloric acid, water, brine,dried and concentrated to afford 2.82g of a pale oil, identified as2(S)-methyl-3-(thiomethyl)propionic acid t-butyl ester.

Part C: To a solution of 2.82 g of 2(S)-methyl-3-(thiomethyl)propionicacid t-butyl ester in 50 mL of acetic acid was added 5.58 g of sodiumperborate and the mixture heated to 55° C. for 17 hours. The reactionwas poured into water, extracted with methylene chloride, washed withaqueous sodium bicarbonate, dried and concentrated to afford 2.68 g of2(S)-methyl-3-(methylsulfonyl)propionic acid t-butyl ester as a whitesolid.

Part D: To 2.68 g of 2(S)-methyl-3-(methylsulfonyl)-propionic acidt-butyl ester was added 20 mL of 4N hydrochloric acid/dioxane and themixture stirred at room temperature for 19 hours. The solvent wasremoved under reduced pressure to afford 2.18 g of crude product, whichwas recrystallized from ethyl acetate/hexane to yield 1.44 g of2(S)-methyl-3-(methylsulfonyl)propionic acid as white crystals.

EXAMPLE 29

Preparation of [1S-[1R*(R*),2S*]]-N-[2-hydroxy-3-[(2-methylpropyl)(3,4-dimethoxyphenylsulfonyl)amino]-1-(phenylmethyl)propyl]-2-methyl-3-(methylsulfonyl)propanamide

Part A: A solution ofN-benzyloxycarbonyl-3(S)-amino-1,2-(S)-epoxy-4-phenylbutane (50.0 g,0.168 mol) and isobutylamine (246 g, 3.24 mol, 20 equivalents) in 650 mLof isopropyl alcohol was heated to reflux for 1.25 hours. The solutionwas cooled to room temperature, concentrated in vacuo and then pouredinto 1 L of stirring hexane whereupon the product crystallized fromsolution. The product was isolated by filtration and air dried to give57.56 g, 92% ofN-[3(S)-benzyloxycarbonylamino-2(R)-hydroxy-4-phenyl]N-isobutylamine, mp108.0–109.5° C., MH+ m/z=371.

Part B: A solution ofN-(3(S)-benzyloxycarbonylamino-2(R)-hydroxy-4-phenyl)N-isobutylamine(1.5356 g, 4.14 mmol) and triethylamine (522 mg, 5.17 mmol) in 15 mL ofdichloromethane was treated with 3,4-dimethoxybenzenesulfonyl chloride(1.0087 g, 4.26 mmol) at room temperature for 14 h. The solvent wasremoved in vacuo and the residue taken up in ethyl acetate and thenwashed with 1N KHSO₄, saturated aqueous NaHCO₃, brine, dried over anhydMgSO₄, filtered and concentrated to give 2.147 g, 90.5%, of a whitesolid, mp 124–127° C., HRFAB MS; M+Li; calc'd. for C₃₀H₃₈N₂O₇S+Li:577.2560. Found: 577.2604.

Part C: A solution of carbamic acid, product from Part B (513 mg, 0.90mmol) in 30 mL of methanol was stirred with 20 mg of palladium blackcatalyst and 10 mL of formic acid for 15 h at room temperature. Thecatalyst was removed by filtration through diatomaceous earth and thefiltrate concentrated in vacuo and the residue taken up in ethylacetate. The ethyl acetate solution was washed with saturated aqueousNaHCO₃, brine and dried over anhyd MgSO₄, filtered and concentrated invacuo to give a white solid, 386 mg, 98%, mp 123–130° C., FAB MS;M+Li⁺=443, that was used directly in the next step without furtherpurification.

Part D: A mixture of 2(S)-methyl-3-methylsulfonyl propionic acid (128mg, 0.77 mmol), N-hydroxybenzotriazole (179.9 mg, 1.17 mmol), and1-(3-dimethylaminopropyl)-3-ethylcarbodiimide hydrochloride (EDC) (177.3mg, 0.92 mmol) was dissolved in 1.5 mL of dimethylformamide (DMF) andallowed to react for 30 min at 0° C.; The amine from Part C (359 mg,0.82 mmol) dissolved in 1 mL of DMF was added to the above mixture andstirred at room temperature for 48 h. The solution was then poured into75 mL of saturated aqueous NaHCO₃ and extracted with ethyl acetate. Theethyl acetate extracts were washed with 5% aqueous citric acid,saturated aqueous NaHCO₃, brine, dried over anhyd MgSO₄, filtered andconcentrated to give a clear oil, 220 mg. The material was crystallizedfrom hexanes and ethyl acetate to give 178 mg, 40% of pure product withmp 130–133° C. HRFAB MS;M+Li⁺; calc'd. for C₂₇H₄₀N₂O₈S₂Li: 591.2386.Found: 591.2396.

EXAMPLE 30

Preparation of [1S-[1R*(R*),2S*]]-N-[2-hydroxy-3-[(3-methylbutyl)(4-aminophenylsulfonyl)amino]-1-(phenylmethyl)propyl]-2-methyl-3-(methylsulfonyl)propanamide

Part A: A solution ofN-benzyloxycarbonyl-3(S)-amino-1,2(S)epoxy-4-phenylbutane (11.54 g,38.81 mmol) and isoamylamine (66.90 g, 0.767 mol, 19.9 equivalents) in90 mL of isopropyl alcohol was-heated to reflux for 3.1 h. The solutionwas cooled to room temperature and partially concentrated in vacuo andthe remaining solution poured into 200 mL of stirring hexanes whereuponthe product crystallized from solution. The product was isolated byfiltration and air dried to give 11.76 g, 79% ofN-[[3(S)-phenylmethoxy)carbonyl)amino-2(R)-hydroxy-4-phenylbutyl]N-[(3-methylbutyl)]amine,mp 118–122° C., FAB MS: MH⁺=385.

Part B: A solution ofN-[[3(S)-(phenylmethoxycarbonyl)amino-2(R)-hydroxy-4-phenylbutyl]N-[(3-methylbutyl)]amine(1.1812 g, 3.07 mmol) and triethylamine (325.7 mg, 3.22 mmol) in 20 mLof dichloromethane was treated with 4-nitrobenzensulfonyl chloride (767mg, 90% purity 3.11 mmol) at room temperature for 10 min. The solventwas removed in vacuo and the residue taken up in ethyl acetate and thenwashed with 1N KHSO₄, saturated aqueous NaHCO₃, brine, dried over anhydMgSO₄, filtered and concentrated to give 2.3230 g, of a tan solid, thatwas crystallized from ethyl acetate and petroleum ether to provide 870mg, 50%, mp 130–132° C. of pure product, HRFAB MS; M+Li, calc'd. forC₂₉H₃₅N₃O₇SLi: 576.2316. Found: 576.2350.

Part C: A solution of product from Part B (574 mg, 1.01 mmol) in 40 mLof methanol, (the solution was not completely homogeneous), was treatedwith 70 mg of 10% palladium on carbon catalyst and hydrogenated at 42psig for 15 h at room temperature. The catalyst was removed byfiltration through diatomaceous earth and the filtrate concentrated invacuo to give a white solid that was crystallized from chloroform, mp123–127° C., FAB MS; M+Li⁺=412, 400 mg, 91%, that was used directly inthe 25 next step without further purification.

Part D: A mixture of 2(S)-methyl-3-methylsulfonyl propionic acid (112.3mg, 0.675 mmol), N-hydroxybenzotriazole (159.1 mg, 1.04 mmol), and1-(3-dimethylaminopropyl)-3-ethylcarbodiimide hydrochloride (EDC) (147.8mg, 0.77 mmol) was dissolved in 1.0 mL of dimethylformamide (DMF) andallowed to react for 30 min at 0° C. The amine from Part C (261.9 mg,0.646 mmol) dissolved in 0.5 mL of DMF was added to the above mixtureand stirred at room temperature for 16.5 h. The solution was then pouredinto 75 mL of saturated aqueous NaHCO₃ and extracted with ethyl acetate.The ethyl acetate extracts were washed with 5% aqueous citric acid,saturated aqueous NaHCO₃, brine, dried over anhyd MgSO₄, filtered andconcentrated to give a white foam, 326.3 mg. The material was purifiedby flash chromatography over silica gel eluting with ethyl acetate toprovide 213.6 mg, 64% of pure product as a white foam, FAB Mg; MH⁺=554.

EXAMPLE 31

Preparation of [1S-[1R*(R*),2S*]]-N-[2-hydroxy-3-[(3-methylbutyl)(4-methoxyphenylsulfonyl)amino]-1-(phenylmethyl)propyl]-2-methyl-3-(methylsulfonyl)propanamide

Part A: A solution ofN-benzyloxycarbonyl-3(S)-amino-1,2(S)epoxy-4-phenylbutane (11.54 g,38.81 mmol) and isoamylamine (66.90 g, 0.767 mol, 19.9 equivalents) in90 mL of isopropyl alcohol was heated to reflux for 3.1 h. The solutionwas cooled to room temperature and partially concentrated in vacuo andthe remaining solution poured into 200 mL of stirring hexanes whereuponthe product crystallized from solution. The product was isolated byfiltration and air dried to give 11.76 g, 79% ofN-[[3(S)-phenylmethoxy)carbonyl)amino-2(R)-hydroxy-4-phenylbutyl]N-[(3-methylbutyl)]amine,mp 118–122° C., FAB MS: MH⁺=385.

Part B: A solution ofN-[[3(S)-phenylmethoxy)carbonyl)amino-2(R)-hydroxy-4-phenylbutyl]N-[(3-methylbutyl)]amine(1.1515 g, 2–99 mmol), and triethylamine (313.5 mg, 3.10 mmol) in 15 mLof dichloromethane was treated with 4-methoxybenzenesulfonyl chloride(630.6 mg, 3.05 mmol) via syringe. The solution was stirred at roomtemperature for 40 min and then concentrated in vacuo. The residue wasdissolved in ethyl acetate and washed with 1N KHSO₄, saturated aqueousNaHCO₃, brine, dried over anhyd MgSO₄, filtered and concentrated to give1.5622 g, of a white foam. The crude product was purified byrecrystallization from a mixture of hexanes and ethyl acetate to give1.1047 g, 67% of pure product mp 95–98° C. High resolution FAB Massspectrum calc'd. for C₃₀H₃₈N₂O₆S: 555.2529. Found: 555.2559.

Part c: A solution of the product from Part B (970 mg, 1.68 mmol) in 30mL of methanol was treated with 70 mg of 10% palladium on carboncatalyst and hydrogenated at 41 psig for 16 h at room temperature. Thecatalyst was removed by filtration and the filtrate concentrated invacuo to give a clear oil that solidified upon standing, mp 81–85° C.,FAB MS; MH⁺=421, 764.1 mg that was used directly in the next step.

Part D: A mixture of 2(S)-methyl-3-methylsulfonyl propionic acid (194mg, 1.17 mmol), N-hydroxybenzotriazole (276 mg, 1.34 mmol), and1-(3-dimethylaminopropyl)-3-ethylcarbodiimide hydrochloride (EDC) (256mg, 1.34 mmol) was dissolved in 3.5 mL of dimethylformamide (DMF) andallowed to react for 30 min at 0° C. The amine from Part C (451.1 mg,1.07 mmol) dissolved in 1.5 mL of DMF was added to the above mixture andstirred at room temperature for 16 h. The solution was then poured into20 mL of saturated aqueous NaHCO₃ and extracted 4 times with ethylacetate. The combined ethyl acetate extracts were washed with 5% aqueouscitric acid, saturated aqueous NaHCO₃, brine, dried over anhyd MgSO₄,filtered and concentrated to give a clear oil that crystallized uponstanding. The material was recrystallized from hexanes and ethyl acetateto give 517.6 mg, 85% of pure product with mp 125–129° C. HRFAB MS;calc'd. for C₂₇H₄₀N₂O₇S₂: 569.2355. Found: 5 569.2397.

EXAMPLE 32

Preparation of [1S-[1R*(R*),2S*]]-N-[2-hydroxy-3-[(2-methylpropyl)(4-methoxyphenylsulfonyl)amino]-1-(phenylmethyl)propyl]-2-methyl-3-(methylsulfonyl)propanamide

Part A: A solution ofN-benzyloxycarbonyl-3(S)-amino-1,2-(S)-epoxy-4-phenylbutane (50.0 g,0.168 mol) and isobutylamine (246 g, 3.24 mol, 20 equivalents) in 650 mLof isopropyl alcohol was heated to reflux for 1.25 hours. The solutionwas cooled to room temperature, concentrated in vacuo and then pouredinto 1 L of stirring hexane whereupon the product crystallized fromsolution. The product was isolated by filtration and air dried to give57.56 g, 92% ofN-[3(S)-benzyloxycarbonylamino-2(R)-hydroxy-4-phenyl]N-isobutylamine, mp108.0–109.5° C., MH+ m/z=371.

Part B:N-(3(S)-benzyloxycarbonylamino-2(R)-hydroxy-4-phenyl]-N-isobutylamine(1.1131 g, 3.00 mmol) and triethylamine (324.0 mg, 3.20 mmol) in 20 mLof dichloromethane was treated with 4-methoxy-benzenesulfonyl chloride(715.4 mg, 3.46 mmol). The solution was stirred at room temperature for6 h and then was concentrated in vacuo. The residue was dissolved inethyl acetate and washed with 1N KHSO₄, saturated aqueous NaHCO₃, brine,dried over anhyd MgSO₄, filtered, and concentrated to give a clear oil.The oil was crystallized from ether to give a white solid 1.273 g, 78%,mp 97–101° C., of pure product, FAB MS; MH⁺=541.

Part C: The product from Part B (930 mg, 1.68 mmol) was dissolved in 30mL of methanol and hydrogenated at 40 psig over 70 mg of 10% palladiumon carbon at room temperature for 17 h. The catalyst was removed byfiltration through diatomaceous earth and the filtrate was concentratedin vacuo to give 704 mg of a clear oil, that solidified upon standing,mp 105–110° C., FAB MS, MH⁺=407, and was used directly in the next stepwithout further purification.

Part D: A mixture of 2-methyl-3(methylsulfonyl)propionic acid (174.9 mg,1.05 mmol), N-hydroxybenzotriazole (230 mg, 1.50 mmol) and EDC (220.5mg, 1.15 mmol) in 2 mL of DMF was stirred at 0° C. for 0.5 mL and thentreated with the amine from Part C (401.2 mg, 0.99 mmol) in 1 mL of DMF.The solution was stirred at room temperature for 16 h and then pouredinto 20 mL of saturated aqueous NaHCO₃. The aqueous solution wasextracted with ethyl acetate and then the ethyl acetate solution waswashed with 5% aqueous citric acid, saturated aqueous NaHCO₃, brine,dried over anhyd MgSO₄, filtered and concentrated in vacuo to give aclear oil, 260 mg, which was purified by flash chromatography on Silicagel eluting with hexanes and ethyl acetate to provide 52.7 mg, 9.6%, mp87–92° C., HRFAB MS; Calc'd for C₂₆H₃₈N₂O₇S₂: 555.2199. Found: 555.2234.

EXAMPLE 33

Preparation of [1S-[1R*(R*),2S*]]-N-[2-hydroxy-3-[(butyl)(4-methoxyphenylsulfonyl)amino]-1-(phenylmethyl)propyl]-2-methyl-3-(methylsulfonyl)propanamide

Part A: From the reaction of (1.48 g, 5.0 mmol) of N-benzyloxycarbonyl3(S)-amino-1,2-(S)-epoxy-4-phenylbutane and (7.314 g, 100.0 mmol) ofn-butylamine, one obtains 1.50 g (80%) ofN-[3(S)-benzyloxycarbonylamino-2(R)-hydroxy-4-phenylbutyl]-N-butylamine,mp 125–128° C., FAB MS, Spectrum: MH⁺=371.

Part B: The amine from Part A (1.52 mg, 4.10 mmol) and triethylamine(488 mg, 4.82 mmol) in 30 mL of dichloromethane was treated with4-methoxybenzenesulfonyl chloride (869 mg, 4.20 mmol) at roomtemperature for 3 h. The solution was removed in vacuo and the residuewas taken up in ethyl acetate. The ethyl acetate solution was washedwith 1N KHSO₄, saturated aqueous NaHCO₃, brine, dried over anhyd MgSO₄,filtered and concentrated to give a white solid that was washed withether and air dried to provide 1.71 g, 77%, mp 118–120° C., FAB MS;M+Li=547, of pure product.

Part C: The product from Part B (1.514 g, 2.80 mmol) in 30 mL ofmethanol was hydrogenated at 40 psig over 110 mg of 10% palladium oncarbon for 16 h at room temperature. The catalyst was removed byfiltration through diatomaceous earth and the filtrate concentrated togive a white solid, 1.20 g, 100%, mp 103–108° C., HRFAB MS; Calc'd forC₂₁H₃₀N₂O₄S: 413.2086. Found: 413.2121, which was used directly in thenext step without further purification.

Part D: A mixture of 2(S)-methyl-3-(methylsulfonyl)propionic acid (354.4mg, 2.13 mmol), N-hydroxybenzotriazole (473.4 mg, 3.09 mmol) and EDC(445.3 mg, 2.33 mmol) in 1.5 mL of DMF was stirred at 0° C. for 25 min.and then treated with the amine from Part C (815 mg, 2.00 mmol) in 2 mLof DMF. The mixture was stirred at room temperature for 16 h and thenpoured into 50 mL of saturated aqueous NaHCO₃ and then extracted withethyl acetate. The ethyl acetate solution was washed with 5% aqueouscitric acid, saturated aqueous NaHCO₃, brine, dried over anhyd MgSO₄,filtered and concentrated in vacuo to give 905 mg of a white foam. Theproduct was purified by flash chromatography on Silica gel eluting withethyl acetate/hexanes to provide 711.6 mg, 65%, of pure product, mp87–92° C., HRFAB MS, M+Li; Calc'd for C₂₆H₃₈N₂O₇S₂Li: 561.2281 Found:561.2346.

EXAMPLE 34

Preparation of [1S-[1R*(R*),2S*]]-N-[2-hydroxy-3-[(propyl)(4-methoxyphenylsulfonyl)amino]-1-(phenylmethyl)propyl]-2-methyl-3-(methylsulfonyl)propanamide

Part A: A solution of N-benzyloxycarbonyl3(S)-amino-1,2-(S)-epoxy-4-phenylbutane (6.06 g, 20.4 mmol) andn-propylamine (20.9 g, 0.35 mmol) in 100 mL of isopropyl alcohol washeated to reflux for 3 h. The solution was then concentrated in vacuo togive a solid that was crystallized from hexanes and ethyl acetate togive 6.53 g, 90%, of the desired product, mp 120–123° C., FAB MS:MH⁺=357.

Part B: A solution of the product from Part A (620 mg, 1.74 mmol) andtriethylamine (250 mg, 2.47 mmol) in 15 mL of dichloromethane wastreated with 4-methoxybenzenesulfonyl chloride (371 mg, 1.79 mmol) atroom temperature for 2.33 h. The solvent was removed in vacuo and theresidue taken up in ethyl acetate and then washed with 1N KHSO₄,saturated aqueous NaHCO₃, brine, dried over anhyd MgSO₄, filtered andconcentrated to give 1.0622 g, of a white foam. The crude product waspurified by flash chromatography over silica gel eluting with hexanesand ethyl acetate to give 615 mg, 67%, of pure product with mp 88–92°C., HRFAB MS; calc'd. for C₂₈H₃₄N₂O₆S: 533.2298. Found: 533.2329.

Part C: A solution of carbamic acid, product from Part B (519 mg, 0.98mmol) in 30 mL of methanol was treated with 70 mg of 10% palladium oncarbon catalyst and hydrogenated at 46 psig for 22 h at roomtemperature. The catalyst was removed by filtration through diatomaceousearth and the filtrate concentrated in vacuo to give a clear oil thatsolidified upon standing, mp 124–127° C., FAB MS; M+Li⁺=399, 387 mg,100%, that was used directly in the next step.

Part D: A mixture of 2(S)-methyl-3-methylsulfonyl propionic acid (138.5mg, 0.83 mmol), N-hydroxybenzotriazole (174.6 mg, 1.14 mmol), and1-(3-dimethylaminopropyl)-3-ethylcarbodiimide hydrochloride (EDC) (171.8mg, 0.90 mmol) was dissolved in 2.5 mL of dimethylformamide (DMF) andallowed to react for 30 min at 0° C. The amine from Part C (304.9 mg,0.78 mmol) dissolved in 1.5 mL of DMF was added to the above mixture andstirred at room temperature for 14.5 h. The solution was then pouredinto 20 mL of saturated aqueous NaHCO₃ and extracted with ethyl acetate.The ethyl acetate extracts were washed with 5% aqueous citric acid,saturated aqueous NaHCO₃, brine, dried over anhyd MgSO₄, filtered andconcentrated to give a white solid. The material was recrystallized fromhexanes and ethyl acetate to give 228 mg, 54% of pure product with mp115–118° C. HRFAB MS; calc'd. for C₂₇H₄₀N₂O₇S₂: 541.2042. Found:541.2064.

EXAMPLE 35

Preparation of Carbamic acid,2R-hydroxy-3-[[(2-aminobenzothiazol-6-yl)sulfonyl](2-methylpropyl)amino]-1S-(phenylmethyl)propyl-,phenylmethyl ester

Carbamic acid,2R-hydroxy-3-[[(4-aminophenyl)sulfonyl](2-methylpropyl)amino]-1S-(phenylmethyl)propyl-,phenylmethyl ester 0.30 g (0.571 mmol) was added to a well mixed powderof anhydrous copper sulfate (1.20 g) and potassium thiocyanate (1.50 g)followed by dry methanol (6 mL) and the resulting black-brown suspensionwas heated at reflux for 2 hrs. The reaction mixture was filtered andthe filtrate was diluted with water (5 mL) and heated at reflux. Ethanolwas added to the reaction mixture, cooled and filtered. The filtrateupon concentration afforded a residue which was chromatographed (ethylacetate:hexane 80:20) to afford 0.26 g (78%) of the desired compound asa solid.

EXAMPLE 36

Preparation of Carbamic acid,2R-hydroxy-3-[[(benzothiazol-6-yl)sulfonyl](2-methylpropyl)amino]-1S-(phenylmethyl)propyl-,phenylmethyl ester

Method 1:

Carbamic acid,2R-hydroxy-3-[[(2-aminobenzothiazol-6-yl)sulfonyl](2-methylpropyl)amino]-1S-(phenylmethyl)propyl-,phenylmethyl ester (0.25 g, 0.429 mmol) was added to a solution ofisoamylnitrite (0.116 mL, 0.858 mmol) in dioxane (5 mL) and the mixturewas heated at 85° C. After the cessation of evolution of nitrogen, thereaction mixture was concentrated and the residue was purified bychromatography (hexane:ethyl acetate 5:3) to afford 0.130 g (53%) of thedesired product as a solid.

Method 2:

Crude benzothiazole-6-sulfonyl chloride in ethyl acetate (100 mL) wasadded to N-[3S-benzyloxycarbonylamino-2R-hydroxy-4-phenyl]-N-isobutylamine (1.03 g, 2.78 mmol) followedby N-methylmorpholine (4 mL). After stirring at room temperature for 18hr, the reaction mixture was diluted with ethyl acetate (100 mL), washedwith citric acid (5%, 100 mL), sodium bicarbonate (saturated, 100 mL)and brine (100 mL), dried (MgSO₄) and concentrated in vacuo. The residuewas chromatographed (silica gel, ethyl acetate: hexane 1:1) to afford0.340 g (23%) of desired product.

EXAMPLE 37

Preparation of Carbamic acid,2R-hydroxy-3-[[(2-aminobenzothiazol-5-yl)sulfonyl](2-methylpropyl)amino]-1S-(phenylmethyl)propyl-,phenylmethyl ester; and Carbamic acid,2R-hydroxy-3-[[(2-aminobenzothiazol-7-yl)sulfonyl](2-methylpropyl)amino]-1S-(phenylmethyl)propyl-,phenylmethyl ester

The carbamic acid,2R-hydroxy-3-[(3-aminophenylsulfonyl)(2-methylpropyl)amino]-1S-(phenylmethyl)propyl-,phenylmethyl ester 0.36 g (0.685 mmol) was added to a well mixed powderof anhydrous copper sulfate (1.44 g) and potassium thiocyanate (1.80 g)followed by dry methanol (10 mL) and the rsulting black-brown suspensionwas heated at reflux for 2 hrs. The reaction mixture was filtered andthe filtrate was diluted with water (5 mL) and heated at reflux. Ethanolwas added to the reaction mixture, cooled and filtered. The filtrateupon concentration afforded a rseidue which was chromatographed (ethylacetate:hexane 1:1) to afford 0.18 g (45%) of the 7-isomer as a solid.Further elution of the column with (ethyl acetate:hexane 3:2) afforded0.80 g (20%) afforded the 5-isomer as a solid.

EXAMPLE 38

Preparation of3S-amino-1-[N-(2-methylpropyl)-N-(4-methoxyphenylsulfonyl)amino]-4-phenyl-2R-butanol

Part A: N-benzyloxycarbonyl-3(S)-amino-1-chloro-4-phenyl-2(S)-butanol

To a solution of N-benzyloxycarbonyl-L-phenylalanine chloromethyl ketone(75 g, 0.2 mol) in a mixture of 800 mL of methanol and 800 mL oftetrahydrofuran was added sodium borohydride (13.17 g, 0.348 mol, 1.54equiv.) over 100 min. The solution was stirred at room temperature for 2hours and then concentrated in vacuo. The residue was dissolved in 1000mL of ethyl acetate and washed with 1N KHSO₄, saturated aqueous NaHCO₃,saturated aqueous NaCl, dried over anhydrous MgSO₄, filtered andconcentrated in vacuo to give an oil. The crude product was dissolved in1000 mL of hexanes at 60° C. and allowed to cool to room temperaturewhere upon crystals formed that were isolated by filtration and washedwith copious amounts of hexanes. This solid was then recrystallized fromhot ethyl acetate and hexanes to provide 32.3 g 43% ofN-benzyloxycarbonyl-3(S)-amino-1-chloro-4-phenyl-2(S)-butanol, mp150–151° C., FAB MS: MLi⁺=340.

Part B: 3(S)-[N-(benzyloxycarbonyl)amino]-1,2(S)-epoxy-4-phenylbutane

A solution of potassium hydroxide (6.52 g. 0.1–16 mol, 1.2 equiv.) in970 mL of absolute ethanol was treated withN-benzyloxycarbonyl-3(S)-amino-1-chloro-4-phenyl-2(S)-butanol (32.3 g,0.097 mol). This solution was stirred at room temperature for 15 minutesand then concentrated in vacuo to give a white solid. The solid wasdissovled in dichloromethane and washed with water, dried over anhydMgSO₄, filetered and concentrated in vacuo to give a white solid. Thesolid was crystallized from hexanes and ethyl acetate to give 22.3 g,77% of 3(S)-[N-(benzyloxycarbonyl)amino]-1,2(S)-epoxy-4-phenylbutane, mp102–103° C., FAB MS: MH⁺=298.

Part C:N-[3(S)-benzyloxycarbonylamino-2(R)-hydroxy-4-phenyl]N-isobutylamine

A solution of N-benzylcarbonyl-3(S)-amino-1,2-(S)-epoxy-4-phenyl butane(50.0 g, 0.168 mol) and isobutylamine (246 g, 3.24 mol, 20 equivalents)in 650 mL of isopropyl alcohol was heated to reflux for 1.25 hours. Thesolution was cooled to room temperature; concentrated in vacuo and thenpoured into 1 L of stirring hexane whereupon the product crystallizedfrom solution. The product was isolated by filtration and air dried togive 57.56 g, 92% ofN[3(S)-benzyloxycarbonylamino-2(R)-hydroxy-4-phenyl]-N-isobutylamine, mp108.0–109.5° C., MH+ m/z=371.

Part D: phenylmethyl[2(R)-hydroxy-3-[N-(2-methylpropyl)-N-(4-methoxyphenylsulfonyl)amino]-1S-(phenylmethyl)propyl]carbamate

The amine from Part C (936.5 mg, 2.53 mmol) and triethylamine (2.88.5mg, 2.85 mmol) was dissolved in 20 mL of dichloromethane and treatedwith 4-methoxybenzenesulfonyl chloride (461 mg, 2.61 mmol). The solutionwas stirred at room temperature for 16 hours and then concentrated invacuo. The residue was dissolved in ethyl acetate and this solution waswashed with 1N KHSO₄, saturated aqueous NaHCO₃, brine, dried over anhydMgSO₄, filtered, and concentrated to give a clear oil 1.234 g. The oilwas crystallized from a mixture of ether and hexanes, 729.3 mg, 56.5% mp95–99° C., FAB MS: MH⁺=511.

Part E:3S-amino-1-[N-(2-methylpropyl)-N-(4-methoxyphenylsulfonyl)amino]-4-phenyl-2R-butanol

A solution of phenylmethyl[2(R)-hydroxy-3-[N-(2-methylpropyl)-N-(4-methoxyphenylsulfonyl)amino]1-S-(phenylmethyl)propylcarbamate (671.1 mg, 1.31 mmol) from Part D in 10 mL of methanol washydrogenated over 50 mg of 10% palladium on carbon at 40 psig at roomtemperature for 15 hours. The catalyst was removed by filtration throughdiatomaceous earth and the filtrate concentrated to give a white foam,474.5 mg, 96%, FAB MS: MH⁺=377.

EXAMPLE 39

Preparation of 1,3-benzodioxole-5-sulfonyl chloride

Method 1:

To a solution of 4.25 g of anhydrous N,N-dimethylformamide at 0° C.under nitrogen was added 7.84 g of sulfuryl chloride, whereupon a solidformed. After stirring for 15 minutes, 6.45 g of 1,3-benzodioxole wasadded, and the mixture heated at 100° C. for 2 hours. The reaction wascooled, poured into ice water, extracted with methylene chloride, driedover magnesium sulfate, filtered and concentrated to give 7–32 g ofcrude material as a black oil. This was chromatographed on silica gelusing 20% methylene chloride/hexane to afford 1.9 g of(1,3-benzodioxol-5-yl)sulfonyl chloride.

Method 2:

To a 22 liter round bottom flask fitted with a mechanical stirrer, acooling condenser, a heating mantle and a pressure equalizing droppingfunnel was added sulfur trioxide DMF complex (2778 g, 18.1 moles).Dichloroethane (4 liters) was then added and stirring initiated.1,3-Benzodioxole (1905 g, 15.6 moles) as then added through the droppingfunnel over a five minute period. The temperature was then raised to 75°C. and held for 22 hours (NMR indicated that the reaction was done after9 hours.) The reaction was cooled to 26° and oxalyl chloride (2290 g,18.1 moles) was added at a rate so as to maintain the temperature below40° C. (1.5 hours). The mixture was heated to 67° C. for 5 hoursfollowed by cooling to 16° C. with an ice bath. The reaction wasquenched with water (51) at a rate which kept the temperature below 20°C. After the addition of water was complete, the mixture was stirred for10 minutes. The layers were separated and the organic layer was washedagain twice with water (51). The organic layer was dried with magnesiumsulfate (500 g) and filtered to remove the drying agent. The solvent wasremoved under vacuum at 50° C. The resulting warm liquid was allowed tocool at which time a solid began to form. After one hour, the solid waswashed with hexane (400 mL), filtered and dried to provide the desiredsulfonyl chloride (2823 g). The hexane wash was concentrated and theresulting solid washed with 400 mL hexane to provide additional sulfonylchloride (464 g). The total yield was 3287 g (95.5% based upon1,3-benzodioxole).

Method 3:

1,4-benzodioxan-6-sulfonyl chloride was prepared according to theprocedure disclosed in EP 583960, incorporated herein by reference.

EXAMPLE 40

Preparation of1-[N-[(1,3-benzodioxol-5-yl)sulfonyl]-N-(2-methylpropyl)amino]-3(S)-[bis(phenylmethyl)amino]-4-phenyl-2(R)-butanol

Method 1:

To a 5000 mL, 3-necked flask fitted with a mechanical stirrer was addedN-[3(S)-[N,N-bis(phenylmethyl)amino]-2(R)-hydroxy-4-phenylbutyl]-N-isobutylamine-oxalicacid salt (354.7 g, 0.7 mole) and 1,4-dioxane (2000 mL). A solution ofpotassium carbonate (241.9 g, 1.75 moles) in water (250 mL) was thenadded. The resultant heterogeneous mixture was stirred for 2 hours atroom temperature followed by the addition of 1,3-benzodioxole-5-sulfonylchloride (162.2 g, 0.735 mole) dissolved in 1,4-dioxane (250 mL) over 15minutes. The reaction mixture was stirred at room temperature for 18hours. Ethyl acetate (1000 mL) and water (500 mL) were charged to thereactor and stirring continued for another 1 hour. The aqueous layer wasseparated and further extracted with ethyl acetate (200 mL). Thecombined ethyl acetate layers were washed with 25% brine solution (500mL) and dried over anhydrous magnesium sulfate. After filtering andwashing the magnesium sulfate with ethyl acetate (200 mL), the solventin the filtrate was removed under reduced pressure yielding the desiredsulfonamide as an viscous yellow foamy oil (440.2 g 105% yield). HPLC/MS(electrospray) (m/z 601 [M+H]⁺].

EXAMPLE 41

Preparation of1-[N-[(1,3-benzodioxol-5-yl)sulfonyl]-N-(2-methylpropyl)amino]-3(S)-amino-4-phenyl-2(R)-butanol.methanesulfonicacid salt

Method 1:

Crude1-[N-[(1,3-benzodioxol-5-yl)sulfonyl]-N-(2-methylpropyl)amino]-3(S)-[bis(phenylmethyl)amino]-4-phenyl-2(R)-butanol(6.2g, 0.010 moles) was dissolved in methanol (40 mL). Methanesulfonicacid (0.969 g, 0.010 moles) and water (5 mL) were then added to thesolution. The mixture was placed in a 500 mL Parr hydrogenation bottlecontaining 20% Pd(OH)₂ on carbon (255 mg, 50% water content). The bottlewas placed in the hydrogenator and purged 5 times with nitrogen and 5times with hydrogen. The reaction was allowed to proceed at 35° C. with63 PSI hydrogen pressure for 18 hours. Additional catalyst (125 mg) wasadded and, after purging, the hydrogenation continued for and additional20 hours. The mixture was filtered through celite which was washed withmethanol (2×10 mL). Approximately one third of the methanol was removedunder reduced pressure. The remaining methanol was removed by aziotropicdistillation with toluene at 80 torr. Toluene was added in 15, 10, 10and 10 mL portions. The product crystallized from the mixture and wasfiltered and washed twice with 10 mL portions of toluene. The solid wasdried at room temperature at 1 torr for 6 hours to yield the amine salt(4.5 g, 84%). HPLC/MS (electrospray) was consistent with the desiredproduct (m/z 421 [M+H]⁺).

Method 2:

Part A:N-[3(S)-[N,N-bis(phenylmethyl)amino]-2(R)-hydroxy-4-phenylbutyl)-N-isobutylamine-oxalicacid salt (2800 g, 5.53 moles) and THF (4 L) were added to a 22 L roundbottom flask fitted with a mechanical stirrer. Potassium carbonate (1921g, 13.9 moles) was dissolved in water (2.8 L) and added to the THFslurry. The mixture was then stirred for one hour.1,3-benzodioxole-5-sulfonyl chloride (1281 g, 5.8 moles) was dissolvedin THF (1.4 L) and added to the reaction mixture over 25 minutes. Anadditional 200 mL of THF was used to rinse the addition funnel. Thereaction was allowed to stir for 14 hours and then water (4 L) wasadded. This mixture was stirred for 30 minutes and the layers allowed toseparate. The layers was removed and the aqueous layer washed twice withTHF (500 mL). The combined THF layers were dried with magnesium sulfate(500 g) for one hour. This solution was then filtered to remove thedrying agent and used in subsequent reactions.

Part B: To the THF solution of crude1-[N-[(1,3-benzodioxol-5-yl)sulfonyl)-N-(2-methylpropyl)amino]-3(S)-[bis(phenylmethyl)amino]-4-phenyl-2(R)-butanolwas added water (500 mL) followed by methane sulfonic acid (531 g, 5.5moles). The solution was stirred to insure complete mixing and added toa 5 gallon autoclave. Pearlman's catalyst (200 g of 20% Pd(OH)₂ on C/50%water) was added to the autoclave with the aid of THF (500 mL). Thereactor was purged four times with nitrogen and four times withhydrogen. The reactor was charged with 60 psig of hydrogen and stirringat 450 rpm started. After 16 hours, HPLC analysis indicated that a smallamount of the mono-benzyl intermediate was still present. Additionalcatalyst (50 g) was added and the reaction was allowed to run overnight.The solution was then filtered through celite (500 g) to remove thecatalyst and concentrated under vacuum in five portions. To eachportion, toluene (500 mL) was added and removed under vacuum toazeotropically removed residual water. The resulting solid was dividedinto three portions and each washed with methyl t-butyl ether (2 L) andfiltered. The residual solvent was removed at room temperature in avacuum oven at less than 1 torr to yield the 2714 g of the expectedsalt.

If desired, the product can be further purified by the followingprocedure. A total of 500 mL of methanol and 170 g of material fromabove was heated to reflux until it all dissolved. The solution wascooled, 200 mL of isopropanol added and then 1000–1300 mL of hexane,whereupon a white solid precipitated. After cooling to 0° C., thisprecipitate was collected and washed with hexane to afford 123 g of thedesired material. Through this procedure, the original material whichwas a 95:5 mixture of alcohol diastereomers was greater than 99:1 of thedesired diastereomer.

EXAMPLE 42

Preparation of2R-hydroxy-3-[[(1,3-benzodioxol-5-yl)sulfonyl](2-methylpropyl)amino]-1S-(phenylmethyl)propylamine

Part A: Preparation of2R-hydroxy-3-[[(1,3-benzodioxol-5-yl)sulfonyl](2-methylpropyl)amino]-1S-(phenylmethyl)propylcarbamicacid phenylmethyl ester

To a solution of 3.19 g(8.6 mmol) of N-[3S-benzyloxycarbonylamino-2R-hydroxy-4-phenyl]-N-isobutylamine in 40 mL of anhydrousmethylene chloride, was added 0.87 g of triethylamine. The solution wascooled to 0° C. and 1.90 g of (1,3-benzodioxol-5-yl)sulfonyl chloridewas added, stirred for 15 minutes at 0° C., then for 17 hours at roomtemperature. Ethyl acetate was added, washed with 5% citric acid,saturated sodium bicarbonate, brine, dried and concentrated to yieldcrude material. This was recrystallized from diethyl ether/hexane toafford 4.77 g of pure2R-hydroxy-3-[[(1,3-benzodioxol-5-yl)sulfonyl](2-methylpropyl)amino]-1S-(phenylmethyl)propylcarbamicacid phenylmethyl ester.

Part B: Preparation of2R-hydroxy-3-[[(1,3-benzodioxol-5-yl)sulfonyl](2-methylpropyl)amino]-1S-(phenylmethyl)propylamine

A solution of 4.11 g of carbamic acid,2R-hydroxy-3-[[(1,3-benzodioxol-5-yl)sulfonyl](2-methylpropyl)amino]-1S-(phenylmethyl)propyl-,phenylmethyl ester in 45 mL of tetrahydrofuran and 25 mL of methanol washydrogenated over 1.1 g of 10% palladium-on-carbon under 50 psig ofhydrogen for 16 hours. The catalyst was removed by filtration and thefiltrate concentrated to afford 1.82 g of the desired2R-hydroxy-3-[[(1,3-benzodioxol-5-yl)sulfonyl](2-methylpropyl)amino]-1S-(phenylmethyl)propylamine.

EXAMPLE 43

Preparation of Benzothiazole-6-sulfonyl Chloride

Part A: Preparation of N-(4-Sulfonamidophenyl)thiourea

A mixture of sulfanilamide (86 g, 0.5 mole), ammonium thiocyanate (76.0g, 0.5 mole) and dilute hydrochloric acid (1.5 N, 1 L) was mechanicallystirred and heated at reflux for 2 hr. About 200 mL of water wasdistilled off and concentration of the reaction mixture afforded asolid. The solid was filtered and was washed with cold water and airdried to afford 67.5 g (59%) of the desired product as a white powder.

Part B: Preparation of 2-Amino-6-sulfonamidobenzothiazole

Bromine (43.20 g, 0.27 mol) in chloroform (200 mL) was added over 1 hr.to a suspension of N-(4-sulfonamidophenyl)-thiourea (27.72, 0.120 mol)in chloroform (800 mL). After the addition, the reaction mixture washeated at reflux for 4.5 hr. The chloroform was removed in vacuo and theresidue was repeatedly distilled with additional amounts of chloroform.The solid obtained was treated with water (600 mL) followed by ammoniumhydroxide (to make it basic), then was heated at reflux for 1 hr. Thecooled reaction mixture was filtered, washed with water and air dried toafford 22.0 g (80%) of the desired product as a white powder.

Part C: Preparation of Benzothiazole-6-sulfonic acid

A suspension of 2-amino-6-sulfonamido-benzothiazole (10.0 g, 43.67 mmol)in dioxane (300 mL) was heated at reflux. Isoamylnitrite (24 mL) wasadded in two portions to the reaction mixture. Vigorous evolution of gaswas observed (the reaction was conducted behind a shield as aprecaution) and after 2 hr., a red precipitate was deposited in thereaction vessel. The reaction mixture was filtered hot, and the solidwas washed with dioxane and was dried. The solid was recrystallized frommethanol-water. A small amount of a precipitate was formed after 2 days.The precipitate was filtered off and the mother liquor was concentratedin vacuo to afford a pale red-orange solid (8.0 g, 85%) of pure product.

Part D: Preparation of 6–Chlorosulfonylbenzothiazole

Thionyl chloride (4 mL) was added to a suspension of thebenzothiazole-6-sulfonic acid (0.60 g, 2.79 mmol) in dichloroethane (15mL) and the reaction mixture was heated at reflux and dimethylformamide(5 mL) was added to the reaction mixture to yield a clear solution.After 1.5 hr. at reflux, the solvent was removed in vacuo and excess HCland thionyl chloride was chased by evaporation with dichloroethane.

EXAMPLE 44 Preparation of2R-hydroxy-3-[[(1,4-benzodioxan-6-yl)sulfonyl](2-methylpropyl)amino]-1S-(phenylmethyl)propylcarbamic acid phenylmethyl ester

To a solution of theN-[3S-[(phenylmethoxycarbonyl)amino]-2R-hydroxy-4-phenylbutyl]-N-(2-methylpropyl)amine(0.5 g, 1.35 mmol) in CH₂Cl₂ (5.0 mL) containing Et3N (0.35 mL, 2.5mmol) was added 1,4-benzodioxan-6-sulfonyl chloride (0.34 g, 1.45 mmol)and stirred at 0° C. for 30 min. After stirring at room temperature for1 hour, the reaction mixture was diluted with CH₂Cl₂ (20 mL), washedwith cold 1N HCl (3×20 mL), water (2×20 mL), satd. NaHCO₃ (2×20 mL) andwater (3×20 mL), dried (Na₂SO₄) and concentrated under reduced pressure.The resulting residue was purified by flash chromatography using 35%EtOAc in hexane to give the desired product as a white amorphous solidwhich crystallized from MeOH as a white powder (0.65 g. 84% yield): m.p. 82–84° C., HRMS-FAB: calcd for C₃₀H₃₇N₂O₇S 569.2321 (MH⁺), found569.2323.

EXAMPLE 45 Preparation of[2R-hydroxy-3-[(benzothiazole-6-sulfonyl)-(2-methylpropyl)amino]-1S-(phenylmethyl)propylaminehydrochloride

Part A: Preparation of[2R-hydroxy-3-[(4-aminophenylsulfonyl)(2-methylpropyl)amino]-1S-(phenylmethyl)propylcarbamicacid t-butyl ester

A mixture of[2R-hydroxy-3-[(4-aminophenylsulfonyl)(2-methylpropyl)-amino]-1S-(phenylmethyl)propylamine3.7 g (9.45 mmol) and BOC-ON (2.33 g, 9.45 mmol) and triethylamine(0.954 g, 9.45 mmol) in tetrahydrofuran (60 mL) was stirred for 16 hoursand concentrated in vacuo. The residue was dissolved in dichloromethane(200 mL), washed with sodium hydroxide (1N, 100 mL) and citric acid (5%,100 mL), dried (MgSO₄), and concentrated to afford 1.18 g (94%) of thedesired product as a white solid.

Part B: Preparation of[2R-Hydroxy-3-[[(2-aminobenzothiazol-6-yl)sulfonyl)(2-methylpropyl)amino]-1S-(phenylmethyl)propylcarbamicacid t-butyl ester

The[2R-hydroxy-3-[(4-aminophenylsulfonyl)(2-methylpropyl)amino]-1S-(phenylmethyl)propylcarbamicacid t-butyl ester (1.12 g, 2.279 mmol) was added to a well mixed powderof anhydrous copper sulfate (4.48 g) and potassium thiocyanate (5.60 g)followed by dry methanol (35 mL) and the resulting black-brownsuspension was heated at reflux for 2 hours. The reaction mixture turnedgrey. The reaction mixture was filtered and the filtrate was dilutedwith water (50 mL) and heated at reflux. Ethanol was added to thereaction mixture, cooled and filtered. The filtrate upon concentrationafforded a residue which was chromatographed (ethyl acetate:methanol90:10) to afford 0.80 g (78%) of the deprotected compound as a solid.This was directly reprotected via the following procedure; (2.25 g,5.005 mmol) BOC-ON (1.24 g), and triethylamine (0.505 g, 5.005 mmol) intetrahydrofuran (20 mL) was stirred at room temperature for 18 hours.The reaction mixture was concentrated and the residue was dissolved indichloromethane (200 mL) and was washed with sodium hydroxide (1N, 100mL) and citric acid (5%, 100 mL) dried (MgSO₄) and concentrated toafford a residue which was chromatographed (ethyl acetate:hexane 3:1) toafford 1.8 g (65%) of the desired product as a solid.

Part C: Preparation of[2R-hydroxy-3-[[(benzothiazol-6-yl)sulfonyl](2-methylpropyl)amino]-1S-(phenylmethyl)propylcarbamicacid t-butyl ester

[2R-Hydroxy-3-[[(2-aminobenzothiazol-6-yl)sulfonyl)(2-methylpropyl)amino]-1S-(phenylmethyl)propylcarbamicacid t-butyl ester (1.80 g, 3.2755 mmol) was added to a solution ofisoamylnitrite (0.88 mL) in dioxane (20 mL) and the mixture was heatedat 85° C. After the cessation of evolution of nitrogen, the reactionmixture was concentrated and the residue was purified by chromatography(hexane:ethyl acetate 1:1) to afford 1.25 g (78%) of the desired productas a solid.

Part D: Preparation of[2R-hydroxy-3-[[(benzothiazol-6-yl)sulfonyl](2-methylpropyl)amino]-1S-(phenylmethyl)propylamine.hydrochloride

[2R-hydroxy-3-[[(benzothiazol-6-yl)sulfonyl)(2-methylpropyl)amino]-1S-(phenylmethyl)propylcarbamicacid t-butyl ester (1.25 g, 2.3385 mmol) was added dioxane/HCl (4N, 10mL) and was stirred at room temperature for 2 hours and concentrated.Excess HCl was chased with toluene to afford 1.0 g (quantitative yield)of the desired product.

EXAMPLE 46

Preparation of 2(S)-methyl-3-(methylsulfonyl)propionic Acid

Part A: To a solution of 200 g (1.23 mol) ofD-(−)-3-acetyl-b-mercaptoisobutyric acid in 1.0 L of methanol, was added161.0 g (2.47 mol) of potassium hydroxide dissolved in 500 mL ofmethanol while maintaining the temperature below 10 C while cooling withan ice bath. After stirring an additional 20 minutes, 117 mL (156 g,1.23 mol) of dimethyl sulfate was added while maintaining thetemperature below 20 C. The ice bath was removed and the mixture stirredfor an additional 60 minutes. The salts were removed by filtration, thesolvents removed under reduced pressure and ethyl acetate added. Afterseparating the aqueous layer, it was acidified with concentratedhydrochloric acid, extracted with ethyl acetate, dried over anhydrousmagnesium sulfate, filtered and concentrated to afford 164 g (99%) ofthe desired 2S-methyl-3-(methylthio)propionic acid, m/e=133 (M−H).

Part B: To a solution of 10.0 g (74.6 mmol) of2S-methyl-3-(methylthio)propionic acid in 150 mL of acetone and 30 mL ofwater, cooled to 18 C in an ice bath, was added 161.8 g (263 mmol) ofOxone. After approximately half of material had been added, thetemperature rose to 24 C, the addition was stopped, temperature loweredto 18 C, then addition continued. After stirring at 15–20 C for 15minutes, the bath was removed and the reaction stirred at roomtemperature for 1 hour. The solids were filtered and washed withacetone, the filtrate concentrated to approximately 40 mL and theresidue dissolved in 200 mL of ethyl acetate. The ethyl acetate layerwas dried with anhydrous magnesium sulfate, filtered and concentrated toafford 11.4 g of an oil. This was dissolved in a minimum of ethylacetate and hexane added to cause a precipitate to form. This wascollected to afford 6.95 g of the desired product, m/e=167 (M+H).

EXAMPLE 47

Preparation ofN-[2R-hydroxy-3-[(2-methylpropyl)[(1,3-benzodioxol-5-yl)sulfonyl]amino]-1S-(phenylmethyl)propyl]-2S-methyl-3-(methylsulfonyl)propanamide

To a solution of 5.0 g (30 mmol) of2S-methyl-3-(methylsulfonyl)propionic acid and 6.90 g (45 mmol)N-hydroxybenzotriazole in 30 mL of anhydrous DMF at 0° C. undernitrogen, was added 6.34 g (33 mmol) of EDC. After approximately 10minutes, the EDC was all dissolved. After 60 minutes at 0° C., asolution of 15–5 g (30 mmol) of2R-hydroxy-3-[[(1,3-benzodioxol-5-yl)sulfonyl)(2-methylpropyl)amino]-1S-(phenylmethyl)propylaminemethanesulfonate in 30 mL of anhydrous DMF, previouly neutralized with3.4 mL (31.6 mmol) of 4-methylmorpholine, was added. After 3 hrs at 0°C., the mixture was then stirred overnight for 17 hrs. The DMF wasremoved under reduced pressure, ethyl acetate added, washed with 5%citric acid, saturated sodium bicarbonate, water, brine, dried overanhydrous magnesium sulfate, filtered and concentrated to afford 16 g ofcrude material, which was 88% pure by HPLC. The product waschromatographed on silica gel using 20%-80% ethyl acetate/hexane toafford the pure product, which was recrystallized from ethylacetate/hexane to afford 8.84 g of pure product, mp 131.8–133.8° C.

Alternatively, to a solution of 35.0 g (211 mmol) of2S-methyl-3-(methylsulfonyl)propionic acid and 48.3g (315 mmol)N-hydroxybenzotriazole in 210 mL of anhydrous DMF at 0° C. undernitrogen, was added 44.4 g (231 mmol) of EDC. After approximately 30minutes, the EDC was all dissolved. After an additional 60 minutes at 0°C., a solution of 108.8 g (211 mmol) of2R-hydroxy-3-[[(1,3-benzodioxol-5-yl)sulfonyl)(2-methylpropyl)amino)-1S-(phenylmethyl)propylaminemethanesulfonate in 350 mL of anhydrous DMF, previouly neutralized with24 mL (22.3 g) of 4-methylmorpholine, was added. After 2 hrs at 0° C.,the mixture was then stirred overnight for 18 hrs. The DMF was removedunder reduced pressure, 1 L of ethyl acetate added, washed with 5%citric acid, saturated sodium bicarbonate, water, brine, dried overanhydrous magnesium sulfate, filtered and concentrated to afford 120.4 gof crude material, which was 90% purity by HPLC. The product wascrystallized twice from 750–1000 mL of absolute ethanol to afford 82.6 gof the desired product, >99% purity by HPLC material.

EXAMPLE 48

Preparation of1-[(2-methylpropyl)[(1,4-benzodioxane-6-yl)sulfonyl]amino]-3S-[(phenylmethoxycarbonyl)amino]-4-phenylbutan-2R-ol

To a solution of theN-[3S-C(phenylmethoxycarbonyl)amino]-2R-hydroxy-4-phenylbutyl]-N-(2-methylpropyl)amine(0.5 g, 1.35 mmol) in CH₂Cl₂ (5.0 mL) containing Et₃N (0.35 mL, 2.5mmol) was added 1,4-benzodioxan-6-sulfonyl chloride (0.34 g, 1.45 mmol)(prepared according to the literature procedure in EP 583960 A2, 1994)and stirred at 0° C. for 30 min. After stirring at room temperature for1 h, the reaction mixture was diluted with CH₂Cl₂ (20 mL), washed withcold 1N HCl (3×20 mL), water (2×20 mL), satd. NaHCO₃ (2×20 mL), water(3×20 mL), dried (Na₂SO₄) and concentrated under reduced pressure. Theresulting residue was purified by flash chromatography using 35% EtOAcin hexane to give the desired 1,4-benzodixan-sulfonamide as a whiteamorphous solid which crystallized from MeOH as a white powder (0.65 g.84%): m.p. 82–84° C., HRMS-FAB: calcd for C₃₀H₃₇N₂O₇S 569.2321 (MH⁺),found 569.2323.

EXAMPLE 49

Preparation ofN-[2R-hydroxy-3-[(2-methylpropyl)[(1,4-benzodioxane-6-yl)sulfonyl]amino]-1S-(phenylmethyl)propyl]-2S-methyl-3-(methylsulfonyl)pronanamide

Part A: A solution of1-[(2-methylpropyl)[(1,4-benzodioxane-6-yl)sulfonyl)amino]-3S-[(phenylmethoxycarbonyl)amino]-4-phenylbutan-2R-ol(0.6 g, 1.06 mmol) in THF (10 mL) was hydrogenated at 50 psi in thepresence of 10% Pd/c (0.4 g, ) for 12 h, at room temperature. Thecatalyst was removed by filtration, filtrate was concentrated underreduced pressure.

Part B: The resulting residue from Part A was dissolved in CH₂Cl₂ (4.0mL) and added to a cooled (0° C.) mixture of2S-methyl-3-(methylsulfonyl)propionic acid (0.2 g, 1.2 mmol), HOBt (0.25g, 1.6 mmol) and EDC (0.24 g, 1.25 mmol) in a solvent mixture of DMF (2mL ) and CH₂Cl₂ (2 mL) and stirred at 0° C. for 2 h. After stirring for3 h at room temperature, the reaction mixture was diluted with CH₂Cl₂(15 mL), washed with cold 0.5N NaOH (2×10 mL), water(3×15 mL), dried(Na₂SO₄) and concentrated in vacuo. The residue was purified by flashchromatography on silica gel using EtOAc as the eluent to furnish thedesired sulfonamide as a white amorphous powder (0.5 g, 82%). R_(t)=19.9min. FABMS m/z 589 (M+Li)⁺; HRMS-FAB calcd. for C₂₇H₃₉N₂O₈S₂ 583.2148(MH)⁺, found 583.2115.

EXAMPLE 50

Preparation ofN-[2R-hydroxy-3-[(2-methylpropyl)[(benzothiazol-5-yl)sulfonyl]amino]-1S-(phenylmethyl)propyl]-2S-methyl-3-(methylsulfonyl)propanamide

A mixture of 2-(S)-methyl-3-methylsulfonylpropionic acid (0.220 g, 1.325mmol), hydroxybenzotriazole (0.178 g, 1.325 mmol), EDC (0.253 g, 1.325mmol) in DMF (20 mL0 was stirred for 1 h at room temperature.[2R-hydroxy-3-[(benzothiazole-6-sulfonyl)-(2-methylpropyl)amino]-1S-(phenylmethyl)propylaminehydrochloride (0.620 g, 1.325 mmol) was added followed by triethylamine(0.260 g, 2.66 mmol) and stirred for 18 h. The reaction mixture wasconcentrated in vacuo and the residue was partitioned between ethylacetate (200 mL) and citric acid (5%, 100 mL). The organic layer waswashed with saturated sodium bicarbonate (100 mL), brine (100 mL) dried(MgSO₄) and concentrated. Chromatography with ethyl acetate: hexane 3:1)afforded 0.330 g (43%) of the desired product as a powder. Calculated:M=581; Found: M+Li=588.

EXAMPLE 51

Preparation ofN-[2R-hydroxy-3-[(2-methylpropyl)[(1,3-benzodioxol-5-yl)sulfonyl]amino]-1S-(phenylmethyl)propyl]-2S-methyl-3-(methylsulfonyl)propanamide

Part A: Preparation ofN-[2R-hydroxy-3-[(2-methylpropyl)[(1,3-benzodioxol-5-yl)sulfonyl]amino]-1S-(phenylmethyl)propyl]-2S-methyl-3-(acetylthio)propanamide

N-Hydroxybenzotriazole (1.79 g, 11.6 mmol) was added to solutionD-(−)-S-acetyl-g-mercaptoisobutyric acid (1.26 g, 7.8 mmol) in 15 mL ofdry dimethylformamide and cooled in an ice bath. To the cooled solution,was added EDC (1.64 g, 8.5 mmol) and stirred for 30 minutes. To this wasadded (3.27 g, 7.8 mmol) of2R-hydroxy-3-[[(1,3-benzodioxol-5-yl)sulfonyl](2-methylpropyl)amino]-1S-(phenylmethyl)propylamineand this was stirred 16 hours with warming to room temperature. Thesolvent was removed and the residue partitioned between ethyl acetateand 5% aqueous potassium hydrogen sulfate. The organic layer was washedwith saturated sodium bicarbonate, and brine, dried over magnesiumsulfate filtered and concentrated to yield 4.4 grams of a crude oil,mass spectrum, m/z=571.8 (M+Li).

Part B: Preparation ofN-[2R-hydroxy-3-[(2-methylpropyl)[(1,3-benzodioxol-5-yl)sulfonyl]amino]-1S-(phenylmethyl)propyl]-2S-methyl-3-mercaptopropanamide

A solution of 4.29 g, (7.8 mmol) of S-acetyl compound from part Adissolved in 100 mL of dry methanol was cooled in an ice bath. Anhydrousammonia was bubbled into the solution for one minute. The solution wasstoppered and stirred to room temperature over 5 hours. The contentswere concentrated on a rotory evaporator, and the residue was dissolvedin ethyl acetate. The organic solution was washed with water, and brine,dried over magnesium sulfate, filtered and concentrated to yield 3.9grams of the free mercaptan which was used without purification.

Part C: Preparation ofN-[2R-hydroxy-3-[(2-methylpropyl)[(1,3-benzodioxol-5-yl)sulfonyl]amino]-1S-(phenylmethyl)propyl)-2S-methyl-3-(methylthio)propanamide

A solution of 1.65 g, (3.15 mmol) of the mercaptan from Part B in 25 mLof tetrahydrofuran was cooled in an ice bath. To this cooled solutionwas added 0.52 g, 3.52 mmol) of DBU followed by 0.22 mL, (3.5 mmoL) ofmethyl iodide and the ice bath was removed after 5 minutes. Afterseveral hours at room temperature the contents were concentrated on arotory evaporator, and the residue was dissolved in ethyl acetate. Theorganics were washed with potassium hydrogen sulfate, sodium bicarbonateand brine, dried over magnesium sulfate, filtered and concentrated toyield 1.48 g of a crude white foam.

Part D: Preparation ofN-[2R-hydroxy-3-[(2-methylpropyl)[(1,3-benzodioxol-5-yl)sulfonyl]amino]-1S-(phenylmethyl)propyl]-2S-methyl-3-(methylsulfonyl)propanamide

To a solution of 440 mg, (0.8 mmol) of thiomethylether from part C abovedissolved in 10 mL of methanol, was added 1.52 g, (24.0 mmol) of oxonefollowed by 10 mL of water. The suspension was stirred at roomtemperature for four hours. The mixture was concentrated on a rototryevaporator, diluted with 50 mL of water and extracted with ethylacetate. The organic layer was washed with sodium bicarbonate, driedover magnesium sulfate, filtered and concentrated to yield 390 mg ofcrude sulfone. Purification by flash chromatography using 1;1 ethylacetate; hexane as the eluant yielded 330 mg of the desired compound;mass spectrum, m/z=575.4 (M+Li).

EXAMPLE 52

Preparation ofN-[2R-hydroxy-3-[(2-methylpropyl)[(1,3-benzodioxol-5-yl)sulfonyl]amino]-1S-(phenylmethyl)propyl]-2S-methyl-3-(methylsulfinyl)propanamide

To a solution ofN-[2R-hydroxy-3-[(2-methylpropyl)[(1,3-benzodioxol-5-yl)sulfonyl)amino]-1S-(phenylmethyl)propyl]-2S-methyl-3-(methylthio)propanamide1.04 g (1.94 mmoL) in 10 mL of glacial acetic acid was added 220 mg(1.94 mmoL) of 30% hydrogen peroxide with stirring. After one hour thereaction was stopped by diluting with water and neutralization by thecareful addition of saturated sodium bicarbonate. The resulting aqueoussuspension was extracted with ethyl acetate, and the organics werewashed with 5% aqueous potassium hydrogen sulfate. The ethyl acetatelayer was dried over magnesium sulfate, filtered, and concentrated toyield a mixture of diasteromeric sulfoxides. Separation of the twodiastereomers by careful flash chromatography yielded 250 mg of fastmoving isomer 1, and 250 mg of slow moving isomer 2, and 400 mg of themixture, mass spectrum, m/z=559.3 (M+Li) isomer 1, and m/z=559.3 isomer2.

EXAMPLE 53

Preparation ofN-[2R-hydroxy-3-[(2-methylpropyl)[(2,3-benzofuran-5-yl)sulfonyl]amino]-1S-(phenylmethyl)propyl]-2S-methyl-3-(methylsulfonyl)propanamide

To a solution of 0.170 g (1 mmol) of2-(S)-methyl-3-methylsulfonylpropionic acid dissolved in 5 mL of drydimethylformamide was added 1.5 equivalents of N-hydroxybenzotriazoleand the solution cooled in an ice bath. To this cooled solution wasadded 0.19 g (1.0 mmol) of EDC and the solution stirred for 30 minutes.To this was added2R-hydroxy-3-[[(2,3-dihydrobenzofuran-5-yl)sulfonyl](2-methylpropyl)amino]-1S-(phenylmethyl)propylamine0.418 g (1.0 mmol) and the reaction stirred for 16 hours. The contentswere concentrated on a rotory evaporator and the residue was dissolvedin ethyl acetate, washed with 5% potassium hydrogen sulfate, saturatedsodium bicarbonate, and brine, dried over magnesium sulfate, filteredand concentrated to yield a crude oil. Purification by flashchromatograpy (SiO2) using an eluant of 1:1 ethyl acetate:hexane yieldedpurified product: mass spectrum, m/z=573.5.

EXAMPLE 54 Preparation of5-chlorosulfonyl-2-carbomethoxyamino-benzimidazole

A solution of 2-carbomethoxyamino-benzimidazole (5.0 g, 0.026 mole) inchlorosulfonic acid (35.00 mL) was stirred at 0° C. for 30 minutes andat room temperature for 3 hours. The resulting dark colored reactionmixture was poured into an ice-water mixture (200 mL), and stirred atroom temperature for 30 minutes. The resulting precipitate was filteredand washed with cold water (500 mL). The solid was dried overnight underhigh vacuum in a desiccator over NaOH pellets to give5-chlorosulfonyl-2-carbomethoxyamino-benzimidazole (5.9 g, 78%) as agrey powder. ¹H NMR (DMSO-d₆) d: 3.89 (s, 3H), 7.55 (d, J=8.4 Hz, 1H),7.65 (d, J=8.4 Hz, 1H), 7.88 (s, 1H). (German Patent DE 3826036)

EXAMPLE 55 Preparation ofN-[2R-hydroxy-3-[N¹-[(2-carbomethoxyamino-benzimidazol-5-yl)sulfonyl]-N¹-(2-methylpropyl)amino]-1S-(phenylmethyl)propyl]carbamicacid phenylmethyl ester

To a cold solution ofN-[3S-[(phenylmethoxycarbonyl)amino]-2R-hydroxy-4-phenylbutyl]-N-(2-methylpropyl)amine(5.0 g, 13.5 mmol) in dichloromethane (70 mL) was added triethylamine(5.95 g, 54.0 mmol) followed by the addition of5-chlorosulfonyl-2-carbomethoxyamino-benzimidazole (4.29 g, 14.85 mmol)in small portions as a solid. The reaction mixture was stirred at 0° C.for 30 minutes and at room temperature for 2.5 hours when reaction ofthe amino alcohol was complete. The mixture was cooled and filtered, andthe filtrate was concentrated. The resulting residue was dissolved inEtOAc (200 mL), washed successively with cold 5% citric acid (3×50 mL),saturated aqueous sodium bicarbonate (3×50 mL) and water (3×100 mL),then dried (Na₂SO₄), concentrated and dried under vacuum. The residuewas triturated with methanol, cooled, filtered, washed with MeOH-EtOAc(1:1, v/v) and dried in a desiccator to give pureN-[2R-hydroxy-3-[[(2-carbomethoxyamino-benzimidazol-5-yl)sulfonyl](2-methylpropyl)-amino]-1S-(phenylmethyl)propyl]carbamicacid phenylmethyl ester (6.02 g, 72%) as a light brown powder: FABMS:m/z=630 (M+Li); HRMS: calcd. for C₃₁H₃₈N₅O₇S (M+H) 624.2492, found624.2488.

EXAMPLE 56 Preparation of2R-hydroxy-3-[[(2-amino-benzimidazol-5-yl)sulfonyl](2-methyl-propyl)amino]-1S-(phenylmethyl)propylamine

A solution ofN-[2R-hydroxy-3-[[(2-carbomethoxyamino-benzimidazol-5-yl)sulfonyl](2-methylpropyl)amino]-1S-(phenylmethyl)propyl]carbamicacid phenylmethyl ester (0.36 g, 0.58 mmol) in 2.5 N methanolic KOH(2.00 mL) was heated at 70° C. under a nitrogen atmosphere for 3 hours.The reaction mixture was diluted with water (10 mL) and extracted withEtOAc (3×15 mL). The combined organic extracts were washed with brine,dried (Na₂SO₄) and concentrated. The resulting residue was purified byreverse-phase HPLC using a 10–90% CH₃CN/H₂O gradient (30 min) at a flowrate of 70 mL/min. The appropriate fractions were combined and freezedried to give pure2R-hydroxy-3-[[(2-amino-benzimidazol-5-yl)sulfonyl](2-methylpropyl)amino]-1S-(phenyl-methyl)propylamine(0.22 g, 58%) as a white powder: FAB-MS m/z=432 (M+H); HRMS: calcd. forC₂₁H₃₀N₅O₃S (M+H) 432.2069, found 432.2071.

EXAMPLE 57 Preparation ofN-[2R-hydroxy-3-[[(2-amino-benzimidazol-5-yl)sulfonyl](2-methylpropyl)-amino]-1S-(phenylmethyl)propyl]carbamicacid phenylmethyl ester

To a solution of2R-hydroxy-3-[[(2-amino-benzimidazol-5-yl)sulfonyl](2-methyl-propyl)amino]-1S-(phenylmethyl)propylamine(0.22 g, 0.33 mmol) in THF (3.00 mL), triethylamine (0.11 g, 1.1 mmol)and benzyloxycarbonyl succinimide (0.09 g, 0.36 mmol) were added, andthe reaction mixture was stirred at room temperature for 16 hours. Thesolution was concentrated, and the residue was partitioned between EtOAc(15 mL) and saturated aqueous sodium bicarbonate. The organic phase waswashed with brine, dried (Na₂SO₄), and concentrated. The resultingresidue was purified by reverse-phase HPLC using a 10–90% CH₃CN/H₂Ogradient (30 min) at a flow rate of 70 mL/min. The appropriate fractionswere combined and freeze dried to give pureN-[2R-hydroxy-3-[[(2-amino-benzimidazol-5-yl)sulfonyl](2-methylpropyl)amino]-1S-(phenylmethyl)propyl]carbamicacid phenylmethyl ester (0.12 g, 61%) as a white powder: FAB-MS m/z=566(M+H); HRMS: calcd. for C₂₉H₃₆N₅O₅S 566.2437 (M+H), found 566.2434.

EXAMPLE 58 Preparation of2R-hydroxy-3-[[(2-carbomethoxyamino-benzimidazol-5-yl)sulfonyl](2-methylpropyl)amino]-1S-(phenylmethyl)propylamine

A solution ofN-[2R-hydroxy-3-[[(2-carbomethoxyamino-benzimidazole-5-yl)sulfonyl)(2-methylpropyl)-amino)-1S-(phenylmethyl)propyl]carbamicacid phenylmethyl ester (2.5 g, 0.4 mmol) in MeOH (10 mL) and THF (50mL) was hydrogenated in the presence of 10% Pd/C (1.2 g) at roomtemperature at 60 psi for 16 hours. The catalyst was removed byfiltration, and the filtrate was concentrated under reduced pressure.The resulting residue was triturated with ether and filtered. The solidsubstance thus obtained was washed with ether and dried in vacuo toafford pure2R-hydroxy-3-[[(2-carbomethoxyamino-benzimidazol-5-yl)sulfonyl](2-methylpropyl)amino]-1S-(phenylmethyl)propylamine(1.5 g, 77%) as an off white powder: R_(t)=12.8 min; FAB-MS m/z=490(M+H); HRMS: calcd. for C₂₃H₃₂N₅O₅S 490.2124 (M+H), found 490.2142.

EXAMPLE 59

Preparation ofN-[2R-hydroxy-3-[(2-methylpropyl)[(2-(carbomethoxyamino)benzimidazol-5-yl)sulfonyl]amino]-1S-(phenylmethyl)propyl]-2S-methyl-3-(methylsulfonyl)propanamide

A mixture of 2-(S)-methyl-3-methylsulfonyl propionic acid (157.0 mg,0.94 mmol), 1-hydroxybenzotriazole hydrate (144.0 mg, 0.94 mmol), and1-(3-dimethylamino-propyl)-3-ethylcarbodiimide hydrochloride (EDC)(180.0 mg, 0.94 mmol) was dissolved in dimethylformamide (5.0 mL), andthe solution was stirred at room temperature for 45 minutes. Then2R-hydroxy-3-[[(2-(carbomethoxyamino)benzimidazol-5-yl)sulfonyl](2-methylpropyl)amino]-1S-(phenylmethyl)propylamine(459 mg, 0.94 mmol) and N-methylmorpholine (202.0 mg, 2.0 mmol) wereadded, and the reaction stirred at room temperature for 16 hours. Thesolution was poured into ethyl acetate (75 mL), and the ethyl acetatelayer was washed with 10% aqueous acetic acid (3×25 mL), saturatedaqueous sodium bicarbonate (3×25 mL) and saturated aqueous sodiumchloride (25 mL). The organic layer was dried over anhydrous sodiumsulfate, and the solvent removed in vacuo. The resulting residue wasdissolved in hot ethyl acetate (25 mL). The solution was cooled to roomtemperature and a precipitate began to form. Hexanes (25 mL) were addedand the solution was stirred at room temperature for 2 hours. Theresulting product was collected by vacuum filtration to give pureN-[2R-hydroxy-3-[(2-methylpropyl)[(2-(carbomethoxyamino)benzimidazol-5-yl)sulfonyl]amino]-1S-(phenylmethyl)propyl]-2S-methyl-3-(methyl-sulfonyl)-propanamideas a white solid (395 mg, 65%); FAB-MS calcd for C₂₈H₃₉N₅O₈S₂ m/z=637(M+H), found m/z=644 (M+Li).

EXAMPLE 60

Preparation ofN-[2R-hydroxy-3-[(2-methylpropyl)[(2-aminobenzothiazol-6-yl)sulfonyl]amino]-1S-(phenylmethyl)propyl]-2S-methyl-3-(methylsulfonyl)propanamide

To a solution of 2-(S)-methyl-3-methylsulfonyl propionic acid (0.249 g,1.5 mmol) in 5 mL of dry dimethylformamide, was added 1.5 equivalents ofN-hydroxy-benzotriazole, and the solution was cooled in an ice bath. Tothis cooled solution was added EDC (0.200 g, 1.5 mmol), and the solutionwas stirred for 30 minutes. To this was added2R-hydroxy-3-[[(2-aminobenzothiazol-6-yl)sulfonyl](2-methyl-propyl)amino]-1S-(phenylmethyl)propylamine(0.673 g, 1.5mmol), and the reaction stirred for 16 hours. The contentswere concentrated in vacuo and the residue was dissolved in ethylacetate, washed with 5% potassium hydrogen sulfate, saturated sodiumbicarbonate, and brine, dried over magnesium sulfate, filtered andconcentrated to yield a crude oil. Purification by flash columnchromatograpy on silica gel using an eluant of 1:1:0.1 ethyl acetate:hexane:methanol yielded pureN-[2R-hydroxy-3-[(2-methylpropyl)[(2-amino-benzothiazol-6-yl)sulfonyl]amino]-1S-(phenylmethyl)propyl]-2S-methyl-3-(methyl-sulfonyl)propanamide;FAB-MS: m/z=598.6 (M+H).

EXAMPLE 61

Preparation ofN-[2R-hydroxy-3-[(2-methylpropyl)[(1,3-benzodioxol-5-yl)sulfonyl]-amino]-1S-(phenylmethyl)propyl]-2S-methyl-3-mercaptopropanamide

Part A: Preparation ofN-[2R-hydroxy-3-[(2-methylpropyl)[(1,3-benzodioxol-5-yl)sulfonyl]amino]-1S-(phenylmethyl)propyl]-2S-methyl-3-(acetylthio)-propanamide

N-Hydroxybenzotriazole (1.64 g, 10.7 mmol) was added to solution ofD-(−)-S-acetyl-β-mercaptoisobutyric acid (1.16 g, 7.1 mmol) in 12 mL ofdry dimethylformamide and cooled in an ice bath. To the cooled solution,was added EDC (1.5 g, 7.8 mmol), and the reaction was then stirred for60 minutes. To this was added (3 g, 7.1 mmol) of2R-hydroxy-3-[[(1,3-benzodioxol-5-yl)sulfonyl](2-methylpropyl)-amino]-1S-(phenylmethyl)propylamine,and this was stirred for 16 hours with warming to room temperature. Thesolvent was removed and the residue partitioned between ethyl acetateand 5% aqueous potassium hydrogen sulfate. The organic layer was washedwith saturated sodium bicarbonate, brine, dried over magnesium sulfate,filtered, and concentrated to yield 3.7 g (91%) of the desired productas a crude oil; mass spectrum, m/z=571.8 (M+Li).

Part B: Preparation ofN-[2R-hydroxy-3-[(2-methylpropyl)[(1,3-benzodioxol-5-yl)sulfonyl]amino]-1S-(phenylmethyl)propyl)-2S-methyl-3-mercaptopropanamide

A solution of 4.29 g, (7.8 mmol) of the S-acetyl compound from Part Adissolved in 100 mL of dry methanol was cooled in an ice bath. Anhydrousammonia was bubbled into the solution for one minute. The solution wasstoppered and stirred at room temperature over 5 hours. The contentswere concentrated on a rotory evaporator, and the residue was dissolvedin ethyl acetate. The organic solution was washed with water, brine,dried over magnesium sulfate, filtered and concentrated to yield 3.9 g(95.6%) of the free mercaptan which was used without furtherpurification; mass spectrum, m/z=529.8 (M+Li).

EXAMPLE 62

Following the procedures of the previous Examples, the compounds setforth in Tables 2 through 8 can be prepared.

TABLE 2

Entry R³ R⁴ 1 isobutyl 2-methyl-1,3-benzodioxol-5-yl 2 isobutyl2-methyl-1,3-benzodioxol-5-yl 3 cyclopentylmethyl2-methyl-1,3-benzodioxol-5-yl 4 cyclohexylmethyl2-methyl-1,3-benzodioxol-5-yl 5 cyclopentylmethyl 1,3-benzodioxol-5-yl 6cyclohexylmethyl 1,3-benzodioxol-5-yl 7 cyclopentylmethylbenzofuran-5-yl 8 cyclohexylmethyl benzofuran-5-yl 0 cyclopentylmethyl2,3-dihydrobenzofuran-5-yl 10 cyclohexylmethyl2,3-dihydrobenzofuran-5-yl 11 isobutyl 1,3-benzodioxol-5-yl 12 isobutylbenzofuran-5-yl 13 isobutyl 2,3-dihydrobenzofuran-5-yl 14 isobutyl1,4-benzodioxan-6-yl 15 isoamyl 1,3-benzodioxol-5-yl 16 isoamyl2,3-dihydrobenzofuran-5-yl 17 isoamyl 1,4-benzodioxan-6-yl 18 isobutylbenzothiazol-6-yl 19 isobutyl 2-amino-benzothiazol-6-yl 20 isobutylbenzoxazol-5-yl 21 cyclopentylmethyl 2,2-difluoro-1,3-benzodioxol-5-yl22 cyclohexylmethyl 2,2-difluoro-1,3-benzodioxol-5-yl

TABLE 3A

Entry R⁵ CH₃— Ph— CH₃CH₂— PhCH₂— CH₃CH₂CH₂— PhCH₂CH₂— CH₃CH₂CH₂CH₂—C₆H₁₁— CH₃CH₂CH₂CH₂CH₂— (CH₃)₂CH₂— CH₂═CHCH₂— 3-propynyl

TABLE 3B

Entry R⁵ CH₃— Ph— CH₃CH₂— PhCH₂— CH₃CH₂CH₂— PhCH₂CH₂— CH₃CH₂CH₂CH₂—C₆H₁₁— CH₃CH₂CH₂CH₂CH₂— (CH₃)₂CH₂ CH₂═CHCH₂— 3-propynyl

TABLE 3C

Entry R⁵ CH₃— Ph— CH₃CH₂— PhCH₂— CH₃CH₂CH₂— PhCH₂CH₂— CH₃CH₂CH₂CH₂—C₆H₁₁— CH₃CH₂CH₂CH₂CH₂— (CH₃)₂CH₂— CH₂═CHCH₂— 3-propynyl

TABLE 3D

Entry R⁵ CH₃— Ph— CH₃CH₂— PhCH₂— CH₃CH₂CH₂— PhCH₂CH₂— CH₃CH₂CH₂CH₂—C₆H₁₁— CH₃CH₂CH₂CH₂CH₂— (CH₃)₂CH₂— CH₂═CHCH₂— 3-propynyl

TABLE 3E

Entry R⁵ CH₃— Ph— CH₃CH₂— PhCH₂— CH₃CH₂CH₂— PhCH₂CH₂— CH₃CH₂CH₂CH₂—C₆H₁₁— CH₃CH₂CH₂CH₂CH₂— (CH₃)₂CH₂— CH₂═CHCH₂— 3-propynyl

TABLE 3F

Entry R⁵ CH₃— Ph— CH₃CH₂— PhCH₂— CH₃CH₂CH₂— PhCH₂CH₂— CH₃CH₂CH₂CH₂—C₆H₁₁— CH₃CH₂CH₂CH₂CH₂— (CH₃)₂CH₂— CH₂═CHCH₂— 3-propynyl

TABLE 3G

Entry R⁵ CH₃— Ph— CH₃CH₂— PhCH₂— CH₃CH₂CH₂— PhCH₂CH₂— CH₃CH₂CH₂CH₂—C₆H₁₁— CH₃CH₂CH₂CH₂CH₂— (CH₃)₂CH₂— CH₂═CHCH₂— 3-propynyl

TABLE 4A

Entry R² (CH₃)₂CHCH₂— (4-FC₆H₅)CH₂— CH₃CH₂CH₂CH₂— (naphth-2-yl)CH₂—CH₃SCH₂CH₂— C₆H₁₁CH₂— C₆H₅CH₂ C₆H₅SCH₂— (4-CH₃OC₆H₅)CH₂—(naphth-2-yl)SCH₂—

TABLE 4B

Entry R⁷ (CH₃)₂CHCH₂— (4-FC₆H₅)CH₂— CH₃CH₂CH₂CH₂— (naphth-2-yl)CH₂—CH₃SCH₂CH₂— C₆H₁₁CH₂— C₆H₅CH₂— C₆H₅SCH₂— (4-CH₃OC₆H₅)CH₂—(naphth-2-yl)SCH₂—

TABLE 4C

Entry R² (CH₃)₂CHCH₂— (4-FC₆H₅)CH₂— CH₃CH₂CH₂CH₂— (naphth-2-yl)CH₂—CH₃SCH₂CH₂— C₆H₁₁CH₂— C₆H₅CH₂— C₆H₅SCH₂— (4-CH₃OC₆H₅)CH₂—(naphth-2-yl)SCH₂—

TABLE 4D

Entry R² (CH₃)₂CHCH₂— (4-FC₆H₅)CH₂— CH₃CH₂CH₂CH₂— (naphth-2-yl)CH₂—CH₃SCH₂CH₂— C₆H₁₁CH₂— C₆H₅CH₂— C₆H₅SCH₂— (4-CH₃OC₆H₅)CH₂—(naphth-2-yl)SCH₂—

TABLE 4E

Entry R² (CH₃)₂CHCH₂— (4-FC₆H₅)CH₂— CH₃CH₂CH₂CH₂— (naphth-2-yl)CH₂—CH₃SCH₂CH₂— C₆H₁₁CH₂— C₆H₅CH₂— C₆H₅SCH₂— (4-CH₃OC₆H₅)CH₂—(naphth-2-yl)SCH₂—

TABLE 4F

Entry R² (CH₃)₂CHCH₂— (4-FC₆H₅)CH₂— CH₃CH₂CH₂CH₂— (naphth-2-yl)CH₂—CH₃SCH₂CH₂— C₆H₁₁CH₂— C₆H₅CH₂— C₆H₅SCH₂— (4-CH₃OC₆H₅)CH₂—(naphth-2-yl)SCH₂—

TABLE 4G

Entry R² (CH₃)₂CHCH₂— (4-FC₆H₅)CH₂— CH₃CH₂CH₂CH₂— (naphth-2-yl)CH₂—CH₃SCH₂CH₂— C₆H₁₁CH₂— C₆H₅CH₂— C₆H₅SCH₂— (4-CH₃OC₆H₅)CH₂—(naphth-2-yl)SCH₂—

TABLE 5A

Entry R³ —CH₂CH₂CH₃—CH₂CH₂CH₂CH₃—CH₂CH(CH₃)₂—CH₂CH₂CH(CH₃)₂

TABLE 5B

Entry R³ —CH₂CH₂CH₃—CH₂CH₂CH₂CH₃—CH₂CH(CH₃)₂—CH₂CH₂CH(CH₃)₂

TABLE 5C

Entry R³ —CH₂CH₂CH₃—CH₂CH₂CH₂CH₃—CH₂CH(CH₃)₂—CH₂CH₂CH(CH₃)₂

TABLE 5D

Entry R³ —CH₂CH₂CH₃—CH₂CH₂CH₂CH₃—CH₂CH(CH₃)₂—CH₂CH₂CH(CH₃)₂

TABLE 5E

Entry R³ —CH₂CH₂CH₃—CH₂CH₂CH₂CH₃—CH₂CH(CH₃)₂—CH₂CH₂CH(CH₃)₂

TABLE 5F

Entry R³ —CH₂CH₂CH₃—CH₂CH₂CH₂CH₃—CH₂CH(CH₃)₂—CH₂CH₂CH(CH₃)₂

TABLE 6A

Entry R¹

TABLE 6B

Entry R¹

TABLE 6C

Entry R¹

TABLE 6D

Entry R¹

TABLE 6E

Entry R¹

TABLE 6F

Entry R¹

TABLE 6G

Entry R¹

TABLE 7

Entry R⁴

TABLE 8A

Entry R⁴

TABLE 8B

Entry R⁴

TABLE 8C

Entry R⁴

TABLE 8D

Entry R⁴

EXAMPLE 62

The compounds of the present invention are effective HIV proteaseinhibitors. Utilizing an enzyme assay as described below, the compoundsset forth in the examples herein disclosed inhibited the HIV enzyme. Thepreferred compounds of the present invention and their calculated IC₅₀(inhibiting concentration 50%, i.e., the concentration at which theinhibitor compound reduces enzyme activity by 50%) values are shown inTable 9. The enzyme method is described below. The substrate is2-Ile-Nle-Phe(p-NO₂)-Gln-ArgNH₂. The positive control is MVT-101(Miller, M. et al, Science, 246, 1149 (1989)] The assay conditions areas follows:

-   -   Assay buffer:        -   20 mM sodium phosphate, pH 6.4        -   20% glycerol        -   1 mM EDTA        -   1 mM DTT        -   0.1% CHAPS            The above described substrate is dissolved in DMSO, then            diluted 10 fold in assay buffer. Final substrate            concentration in the assay is 80 μM. HIV protease is diluted            in the assay buffer to a final enzyme concentration of 12.3            nanomolar, based on a molecular weight of 10,780.

The final concentration of DMSO is 14% and the final concentration ofglycerol is 18%. The test compound is dissolved in DMSO and diluted inDMSO to 10× the test concentration; 10 μl of the enzyme preparation isadded, the materials mixed and then the mixture is incubated at ambienttemperature for 15 minutes. The enzyme reaction is initiated by theaddition of 40 μl of substrate. The increase in fluorescence ismonitored at 4 time points (0, 8, 16 and 24 minutes) at ambienttemperature. Each assay is carried out in duplicate wells.

The preceding examples can be repeated with similar success bysubstituting the generically or specifically described reactants and/oroperating conditions of this invention for those used in the precedingexamples.

EXAMPLE 63

The effectiveness of various compounds were determined in theabove-described enzyme assay and in a CEM cell assay.

The HIV inhibition assay method of acutely infected cells is anautomated tetrazolium based colorimetric assay essentially that reportedby Pauwles et al, J. Virol. Methods, 20, 309–321 (1988). Assays wereperformed in 96-well tissue culture plates. CEM cells, a CD4⁺ cell line,were grown in RPMI-1640 medium (Gibco) supplemented with a 10% fetalcalf serum and were then treated with polybrene (2 μg/ml). An 80 μlvolume of medium containing 1×10⁴ cells was dispensed into each well ofthe tissue culture plate. To each well was added a 100 μl volume of testcompound dissolved in tissue culture medium (or medium without testcompound as a control) to achieve the desired final concentration andthe cells were incubated at 37° C. for 1 hour. A frozen culture of HIV-1was diluted in culture medium to a concentration of 5×10⁴ TCID₅₀ per ml(TCID₅₀=the dose of virus that infects 50% of cells in tissue culture),and a 20 μL volume of the virus sample (containing 1000 TCID₅₀ of virus)was added to wells containing test compound and to wells containing onlymedium (infected control cells). Several wells received culture mediumwithout virus (uninfected control cells). Likewise, the intrinsictoxicity of the test compound was determined by adding medium withoutvirus to several wells containing test compound. In summary, the tissueculture plates contained the following experiments:

Cells Drug Virus 1. + − − 2. + + − 3. + − + 4. + + +

In experiments 2 and 4 the final concentrations of test compounds were1, 10, 100 and 500 μg/ml. Either azidothymidine (AZT) or dideoxyinosine(ddI) was included as a positive drug control. Test compounds weredissolved in DMSO and diluted into tissue culture medium so that thefinal DMSO concentration did not exceed 1.5% in any case. DMSO was addedto all control wells at an appropriate concentration.

Following the addition of virus, cells were incubated at 37° C. in ahumidified, 5% CO₂ atmosphere for 7 days. Test compounds could be addedon days 0, 2 and 5 if desired. On day 7, post-infection, the cells ineach well were resuspended and a 100 μl sample of each cell suspensionwas removed for assay. A 20 μL volume of a 5 mg/ml solution of3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) wasadded to each 100 μL cell suspension, and the cells were incubated for 4hours at 27° C. in a 5% CO₂ environment. During this incubation, MTT ismetabolically reduced by living cells resulting in the production in thecell of a colored formazan product. To each sample was added 100 μl of10% sodium dodecylsulfate in 0.01 N HCl to lyse the cells, and sampleswere incubated overnight. The absorbance at 590 nm was determined foreach sample using a Molecular Devices microplate reader. Absorbancevalues for each set of wells is compared to assess viral controlinfection, uninfected control cell response as well as test compound bycytotoxicity and antiviral efficacy.

TABLE 9 IC₅₀ EC₅₀ Entry Compound (nM) (nM) 1

2 20 2

2 28 3

3 26 4

2 12 5

2 52 isomer 1, Example 52 6

7 80 isomer 2, Example 52

The compounds of the present invention are effective antiviral compoundsand, in particular, are effective retroviral inhibitors as shown above.Thus, the subject compounds are effective HIV protease inhibitors. It iscontemplated that the subject compounds will also inhibit otherretroviruses such as other lentiviruses in particular other strains ofHIV, e.g. HIV-2, human T-cell leukemia virus, respiratory syncitialvirus, simia immunodeficiency virus, feline leukemia virus, felineimmuno-deficiency virus, hepadnavirus, cytomegalovirus and picornavirus.Thus, the subject compounds are effective in the treatment, proplylaxisof retroviral infections and/or the prevention of the spread ofretroviral infections.

The subject compounds are also effective in preventing the growth ofretroviruses in a solution. Both human and animal cell cultures, such asT-lymphocyte cultures, are utilized for a variety of well knownpurposes, such as research and diagnostic procedures includingcalibrators and controls. Prior to and during the growth and storage ofa cell culture, the subject compounds may be added to the cell culturemedium at an effective concentration to prevent the unexpected orundesired replication of a retrovirus that may inadvertently,unknowingly or knowingly be present in the cell culture. The virus maybe present originally in the cell culture, for example HIV is known tobe present in human T-lymphocytes long before it is detectable in blood,or through exposure to the virus. This use of the subject compoundsprevents the unknowing or inadvertent exposure of a potentially lethalretrovirus to a researcher or clinician.

Compounds of the present invention can possess one or more asymmetriccarbon atoms and are thus capable of existing in the form of opticalisomers as well as in the form of racemic or nonracemic mixturesthereof. The optical isomers can be obtained by resolution of theracemic mixtures according to conventional processes, for example byformation of diastereoisomeric salts by treatment with an opticallyactive acid or base. Examples of appropriate acids are tartaric,diacetyltartaric, dibenzoyltartaric, ditoluoyltartaric andcamphorsulfonic acid and then separation of the mixture ofdiastereoisomers by crystallization followed by liberation of theoptically active bases from these salts. A different process forseparation of optical isomers involves the use of a chiralchromatography column optimally chosen to maximize the separation of theenantiomers. Still another available method involves synthesis ofcovalent diastereoisomeric molecules by reacting compounds of Formula Iwith an optically pure acid in an activated form or an optically pureisocyanate. The synthesized diastereoisomers can be separated byconventional means such as chromatography, distillation, crystallizationor sublimation, and then hydrolyzed to deliver the enantiomerically purecompound. The optically active compounds of Formula I can likewise beobtained by utilizing optically active starting materials. These isomersmay be in the form of a free acid, a free base, an ester or a salt.

The compounds of the present invention can be used in the form of saltsderived from inorganic or organic acids. These salts include but are notlimited to the following: acetate, adipate, alginate, citrate,aspartate, benzoate, benzenesulfonate, bisulfate, butyrate, camphorate,camphorsulfonate, digluconate, cyclopentanepropionate, dodecylsulfate,ethanesulfonate, glucoheptanoate, glycerophosphate, hemisulfate,heptanoate, hexanoate, fumarate, hydrochloride, hydrobromide,hydroiodide, 2-hydroxy-ethanesulfonate, lactate, maleate,methanesulfonate, nicotinate, 2-naphthalenesulfonate, oxalate, palmoate,pectinate, persulfate, 3-phenylpropionate, picrate, pivalate,propionate, succinate, tartrate, thiocyanate, tosylate, mesylate andundecanoate. Also, the basic nitrogen-containing groups can bequaternized with such agents as lower alkyl halides, such as methyl,ethyl, propyl, and butyl chloride, bromides, and iodides; dialkylsulfates like dimethyl, diethyl, dibutyl, and diamyl sulfates, longchain halides such as decyl, lauryl, myristyl and stearyl chlorides,bromides and iodides, aralkyl halides like benzyl and phenethylbromides, and others. Water or oil-soluble or dispersible products arethereby obtained.

Examples of acids which may be employed to form pharmaceuticallyacceptable acid addition salts include such inorganic acids ashydrochloric acid, sulphuric acid and phosphoric acid and such organicacids as oxalic acid, maleic acid, succinic acid and citric acid. Otherexamples include salts with alkali metals or alkaline earth metals, suchas sodium, potassium, calcium or magnesium or with organic bases.

Total daily dose administered to a host in single or divided doses maybe in amounts, for example, from 0.001 to 10 mg/kg body weight daily andmore usually 0.01 to 1 mg. Dosage unit compositions may contain suchamounts of submultiples thereof to make up the daily dose.

The amount of active ingredient that may be combined with the carriermaterials to produce a single dosage form will vary depending upon thehost treated and the particular mode of administration.

The dosage regimen for treating a disease condition with the compoundsand/or compositions of this invention is selected in accordance with avariety of factors, including the type, age, weight, sex, diet andmedical condition of the patient, the severity of the disease, the routeof administration, pharmacological considerations such as the activity,efficacy, pharmacokinetic and toxicology profiles of the particularcompound employed, whether a drug delivery system is utilized andwhether the compound is administered as part of a drug combination.Thus, the dosage regimen actually employed may vary widely and thereforemay deviate from the preferred dosage regimen set forth above.

The compounds of the present invention may be administered orally,parenterally, by inhalation spray, rectally, or topically in dosage unitformulations containing conventional nontoxic pharmaceuticallyacceptable carriers, adjuvants, and vehicles as desired. Topicaladministration may also involve the use of transdermal administrationsuch as transdermal patches or iontophoresis devices. The termparenteral as used herein includes subcutaneous injections, intravenous,intramuscular, intrasternal injection, or infusion techniques.

Injectable preparations, for example, sterile injectable aqueous oroleaginous suspensions may be formulated according to the known artusing suitable dispersing or wetting agents and suspending agents. Thesterile injectable preparation may also be a sterile injectable solutionor suspension in a nontoxic parenterally acceptable diluent or solvent,for example, as a solution in 1,3-butanediol. Among the acceptablevehicles and solvents that may be employed are water, Ringer's solution,and isotonic sodium chloride solution. In addition, sterile, fixed oilsare conventionally employed as a solvent or suspending medium. For thispurpose any bland fixed oil may be employed including synthetic mono- ordiglycerides. In addition, fatty acids such as oleic acid find use inthe preparation of injectables.

Suppositories for rectal administration of the drug can be prepared bymixing the drug with a suitable nonirritating excipient such as cocoabutter and polyethylene glycols which are solid at ordinary temperaturesbut liquid at the rectal temperature and will therefore melt in therectum and release the drug.

Solid dosage forms for oral administration may include capsules,tablets, pills, powders, and granules. In such solid dosage forms, theactive compound may be admixed with at least one inert diluent such assucrose lactose or starch. Such dosage forms may also comprise, as innormal practice, additional substances other than inert diluents, e.g.,lubricating agents such as magnesium stearate. In the case of capsules,tablets, and pills, the dosage forms may also comprise buffering agents.Tablets and pills can additionally be prepared with enteric coatings.

Liquid dosage forms for oral administration may include pharmaceuticallyacceptable emulsions, solutions, suspensions, syrups, and elixirscontaining inert diluents commonly used in the art, such as water. Suchcompositions may also comprise adjuvants, such as wetting agents,emulsifying and suspending agents, and sweetening, flavoring, andperfuming agents.

While the compounds of the invention can be administered as the soleactive pharmaceutical agent, they can also be used in combination withone or more immunomodulators, antiviral agents or other antiinfectiveagents. For example, the compounds of the invention can be administeredin combination with AZT, DDI, -DDC or with glucosidase inhibitors, suchas N-butyl-1-deoxynojirimycin or prodrugs thereof, for the prophylaxisand/or treatment of AIDS. When administered as a combination, thetherapeutic agents can be formulated as separate compositions which aregiven at the same time or different times, or the therapeutic agents canbe given as a single composition.

The foregoing is merely illustrative of the invention and is notintended to limit the invention to the disclosed compounds. Variationsand changes which are obvious to one skilled in the art are intended tobe within the scope and nature of the invention which are defined in theappended claims.

From the foregoing description, one skilled in the art can easilyascertain the essential characteristics of this invention, and withoutdeparting from the spirit and scope thereof, can make various changesand modifications of the invention to adapt it to various usages andconditions.

1. Compound represented by the formula:

or a pharmaceutically acceptable salt thereof, wherein t represents 0, 1or 2; R¹ represents hydrogen, alkyl of 1–5 carbon atoms, alkenyl of 2–5carbon atoms, alkynyl of 2–5 carbon atoms, hydroxyalkyl of 1–3 carbonatoms, alkoxyalkyl of 1–3 alkyl and 1–3 alkoxy carbon atoms, cyanoalkylof 1–3 alkyl carbon atoms, —CH₂CONH₂, —CH₂CH₂CONH₂, —CH₂S(O)₂NH₂,—CH₂SCH₃, —CH₂S(O)CH₃ or —CH₂S(O)₂CH₃ radicals; R² represents radicalsof alkyl of 1–5 carbon atoms, aralkyl of 1–3 alkyl carbon atoms,alkylthioalkyl of 1–3 alkyl carbon atoms, arylthioalkyl of 1–3 alkylcarbon atoms or cyloalkylalkyl of 1–3 alkyl carbon atoms and 3–6 ringmember carbon atoms; R³ represents radicals of alkyl radical of 1–5carbon atoms, cyloalkyl of 5–8 ring members or cycloalkylmethyl radicalof 3–6 ring members; R⁴ represents benzo fused 5 to 6 ring memberheteroaryl or benzo fused 5 to 6 ring member heterocyclo radicals, or aradical of the formula

wherein A and B each independently represent O, S, SO or SO₂; R⁶represents deuterium, alkyl of 1–5 carbon atoms, fluoro or chlororadicals; R⁷ represents hydrogen, deuterium, methyl, fluoro or chlororadicals; or a radical of the formula

wherein Z represents O, S or NH; and R⁹ represents a radical of formula

wherein Y represents O, S or NH; X represents a bond, O or NR²¹; R²⁰represents hydrogen, alkyl of 1 to 5 carbon atoms, alkenyl of 2 to 5carbon atoms, alkynyl of 2 to 5 carbon atoms, aralkyl of 1 to 5 alkylcarbon atoms, heteroaralkyl of 5 to 6 ring members and 1 to 5 alkylcarbon atoms, heterocycloalkyl of 5 to 6 ring members and 1 to 5 alkylcarbon atoms, aminoalkyl of 2 to 5 carbon atoms, N-mono-substituted orN,N-disubstituted aminoalkyl of 2 to 5 alkyl carbon atoms wherein saidsubstituents are radicals of alkyl of 1 to 3 carbon atoms, aralkyl of 1to 3 alkyl carbon atoms radicals, carboxyalkyl of 1 to 5 carbon atoms,alkoxycarbonylalkyl of 1 to 5 alkyl carbon atoms, cyanoalkyl of 1 to 5carbon atoms or hydroxyalkyl of 2 to 5 carbon atoms; R²¹ representshydrogen radical or alkyl radical of 1 to 3 carbon atoms; or the radicalof formula —NR²⁰R²¹ represents a 5 to 6 ring member heterocyclo radical;and R²² represents alkyl radical of 1 to 3 carbon atoms or R²⁰R²¹N-alkylradical of 1 to 3 alkyl carbon atoms; and R⁵ represents an alkyl radicalof 1–5 carbon atoms, alkenyl radical of 2–5 carbon atoms, alkynylradical of 2–5 carbon atoms or aryl substituted alkyl radical of 1–5carbon atoms.
 2. Compound of claim 1, or a pharmaceutically acceptablesalt, thereof, wherein t represents 1 or 2; R¹ represents hydrogenradical, alkyl radical of 1–3 carbon atoms, alkenyl radical of 2–3carbon atoms, alkynyl radical of 2–3 carbon atoms radicals orcyanomethyl; R² represents radicals of alkyl of 3–5 carbon atoms,arylmethyl, alkylthioalkyl of 1–3 alkyl carbon atoms, arylthiomethyl orcyloalkylmethyl of 5–6 ring member carbon atoms radicals; R³ representsalkyl of 1–5 carbon atoms, cyloalkylmethyl of 3–6 ring members,cyclohexyl or cycloheptyl radicals; R⁴ represents2-amino-benzothiazol-5-yl, 2-amino-benzothiazol-6-yl, benzothiazol-5-yl,benzothiazol-6-yl, benzoxazol-5-yl, 2,3-dihydrobenzofuran-5-yl,benzofuran-5-yl, 1,3-benzodioxol-5-yl or 1,4-benzodioxan-6-yl radicals;or a radical of the formula

wherein A and B each represent O; R⁶ represents deuterium, methyl,ethyl, propyl, isopropyl or fluoro radicals; and R⁷ represents hydrogen,deuterium, methyl or fluoro radicals; or a radical of the formula

wherein Z represents O, S or NH; and R⁹ represents a radical of formula

wherein Y represents O, S or NH; X represents a bond, O or NR²¹; R²⁰represent hydrogen, alkyl of 1 to 5 carbon atoms, phenylalkyl of 1 to 3alkyl carbon atoms, heterocycloalkyl of 5 to 6 ring members and 1 to 3alkyl carbon atoms, or N-mono-substituted or N,N-disubstitutedaminoalkyl of 2 to 3 alkyl carbon atoms wherein said substituents arealkyl radicals of 1 to 3 carbon atoms; and R²¹ represents hydrogen ormethyl radicals; or the radical of formula —NR²⁰R²¹ representspyrrolidinyl, piperidinyl, piperazinyl, 4-methylpiperazinyl,4-benzylpiperazinyl, morpholinyl or thiamorpholinyl radicals; and R²²represents alkyl radical of 1 to 3 carbon atoms; and R⁵ represents analkyl radical of 1–5 carbon atoms, alkenyl radical of 3–4 carbon atoms,alkynyl radical of 3–4 carbon atoms or aryl substituted alkyl radical of3–4 carbon atoms.
 3. Compound of claim 2, or a pharmaceuticallyacceptable salt thereof, wherein R¹ represents hydrogen, methyl, ethylor cyanomethyl radicals; R² represents isobutyl, n-butyl, CH₃SCH₂CH₂—,phenylthiomethyl, (2-naphthylthio)methyl, benzyl, 4-methoxyphenylmethyl,4-hydroxyphenylmethyl, 4-fluorophenylmethyl or cyclohexylmethylradicals; R³ represents propyl, isoamyl, isobutyl, butyl, cyclohexyl,cycloheptyl, cylopentylmethyl or cyclohexylmethyl radicals; and R⁴represents benzothiazol-5-yl, benzotbiazol-6-yl, benzoxazol-5-yl,2,3-dihydrobenzofuran-5-yl, benzofuran-5-yl, 1,3-benzodioxol-5-yl,2-methyl-1,3-benzodioxol-5-yl, 2,2-dimethyl-1,3-benzodioxol-5-yl,2,2-dideutero-1,3-benzodioxol-5-yl, 2,2-difluoro-1,3-benzodioxol-5-yl or1,4-benzodioxan-6-yl radicals; or a radical of the formula

wherein Z represents O, S or NH; and R⁹ represents a radical of formula

wherein Y represents O, S or NH; X represents a bond, O or NR²¹; R²⁰represents hydrogen, methyl, ethyl propyl, isobutyl, benzyl,2-(1-pyrrolidinyl) ethyl, 2-(1-piperidinyl)ethyl,2-(1-piperazinyl)ethyl, 2-(4-methylpiperazin-1-yl)ethyl,2-(1-morpholinyl)ethyl, 2-(1-thiamorpholinyl)ethyl or2-(N,N-dimethylamino)ethyl radicals; R²¹ represents a hydrogen radical;and R²² represents methyl radical; and R⁵ represents an alkyl radical of1–5 carbon atoms or phenyl substituted alkyl radical of 2–4 carbonatoms.
 4. Compound of claim 3 or a pharmaceutically acceptable saltthereof, wherein R¹ represents methyl or ethyl radicals; R² representsbenzyl, 4-fluorophenylmethyl or cyclo hexylmethyl radicals; R⁴represents benzothiazol-5-yl, benzothiazol-6-yl,2,3-dihydrobenzofuran-5-yl, benzofuran-5-yl, 1,3-benzodioxol-5-yl,2-methyl-1-,3-benzodioxol-5-yl, 2,2-dimethyl-1,3 benzodioxol-5-yl,2,2-dideutero-1,3-benzodioxol-5-yl, 2,2-difluro-1,3-benzodioxol-5-yl,1,4-benzodioxan-6-yl, 2-(methoxycarbonylamino)benzothiazol-6-yl or2-(methoxycarbonylamino)benzimidazol-5-yl radicals; and R⁵ representsmethyl, ethyl, propyl, isopropyl or 2-phenylethyl radicals. 5.Composition comprising a compound of claim 1 or a pharmaceuticallyacceptable salt thereof and a pharmaceutically acceptable carrier. 6.Method of inhibiting a retroviral protease to treat a retroviralinfection or AIDS comprising administering an effective amount of acompound of claim 1 or a pharmaceutically acceptable salt thereof. 7.Method of preventing replication of retrovirus in vitro comprisingadministering an effective amount of a compound of claim 1 or apharmaceutically acceptable salt thereof.